Prostate cancer genes

ABSTRACT

The present invention relates to all facets of novel polynucleotides, the polypeptides they encode, antibodies and specific binding partners thereto, and their applications to research, diagnosis, drug discovery, therapy, clinical medicine, forensic science and medicine, etc. The polynucleotides are differentially-regulated in prostate cancer and are therefore useful in variety of ways. including, but not limited to, as molecular markers, as drug targets, and for detecting, diagnosing, staging, monitoring, prognosticating, preventing or treating, determining predisposition to, etc., diseases and conditions, to prostate cancer.

This application claims the benefit of U.S. Provisional Application Nos.60/331,042 which was filed Nov. 7, 2001, 60/331,041 which was filed Nov.7, 2001, 60/340,251 which was filed Dec. 18, 2001, and 60/344,791 whichwas filed Jan. 7, 2002, which are hereby incorporated by reference intheir entirety.

DESCRIPTION OF THE DRAWINGS

SEQ ID NOS. 1-49 show the nucleotide and amino acid sequences ofdifferentially-regulated genes. The polynucleotides are human cDNAs.

FIG. 1 shows amino acid sequence comparisons between Pc0378 (SEQ ID NO6), NM_(—)017934 (SEQ ID NO 50), and AAG45146 (SEQ ID NO 51).

FIG. 2 shows an amino acid comparison between Pc0099 (SEQ ID NO 39) andXM_(—)047995 (SEQ ID NO 52).

DESCRIPTION OF THE INVENTION

The present invention relates to all facets of novel polynucleotides,the polypeptides they encode, antibodies and specific binding partnersthereto, and their applications to research, diagnosis, drug discovery,therapy, clinical medicine, forensic science and medicine, etc. Thepolynucleotides are differentially regulated in prostate cancer and aretherefore useful in variety of ways, including, but not limited to, asmolecular markers, as drug targets, and for detecting, diagnosing,staging, monitoring, prognosticating, preventing or treating,determining predisposition to, etc., diseases and conditions, especiallyrelating to prostate cancer. The identification of specific genes, andgroups of genes, expressed in pathways physiologically relevant toprostate cancer permits the definition of functional and diseasepathways, and the delineation of targets in these pathways which areuseful in diagnostic, therapeutic, and clinical applications. Thepresent invention also relates to methods of using the polynucleotidesand related products (proteins, antibodies, etc.) in business andcomputer-related methods, e.g., advertising, displaying, offering,selling, etc., such products for sale, commercial use, licensing, etc.

Prostate cancer is the most common form of cancer diagnosed in theAmerican male, occurring predominantly in males over age 50. The numberof men diagnosed with prostate cancer has steadily increased as a resultof the increasing population of older men. The American Cancer Societyestimates that in the year 2000, about 180,000 American men werediagnosed with prostate cancer and about 32,000 died from the disease.In comparison, 1998 estimates for lung cancer in men were 171,500 casesand 160,100 deaths, and for colorectal cancer, the estimates were131,600 cases and 56,000 deaths. Despite these high numbers, 89 percentof men diagnosed with the disease will survive at least five years and63 percent will survive at least 10 years.

Patients having prostate cancer display a wide range of phenotypes. Insome men, following detection, the tumor remains a latent histologicaltumor and does not become clinically significant. However, in other men,the tumor progresses rapidly, metastasizing and killing the patient in arelatively short time. Prostate cancer can be cured if the tumor isconfined to a small region of the gland and is discovered at earlystage. In such cases, radiation or surgical removal often results incomplete elimination of the disease. Frequently, however, the prostatecancer has already spread to surrounding tissue and metastasized toremote locations. In these cases, radiation and other therapies, areless likely to effect a complete cure.

Androgen deprivation is a conventional therapy to treat prostate cancer.Androgen blockade can be achieved through several different routes.Androgen suppressive drugs include, e.g., Lupron (leuprolide acetate),Casodex (bicalutamide), Eulexin (flutamide), Nilandron (nilutamide),Zoladex (goserelin acetate implant), and Viadur (leuprolide acetate),which act through several different mechanisms. While these drugs mayoffer remission and tumor regression in many cases, often thetherapeutic effects are only temporary. Prostate tumors lose theirsensitivity to such treatments, and become androgen-independent. Thus,new therapies are clearly needed.

The first clinical symptoms of prostate cancer are typically urinarydisturbances, including painful and more frequent urination. Diagnosisfor prostate cancer is usually accomplished using a combination ofdifferent procedures. Since the prostate is located next to the rectum,rectal digital examination allows the prostate to be examined manuallyfor the presence of hyperplasia and abnormal tissue masses. Usually,this is the first line of detection. If a palpable mass is observed, ablood specimen can be assayed for prostate-specific antigen (PSA). Verylittle PSA is present in the blood of a healthy individual, but BPH andprostate cancer can cause large amounts of PSA to be released into theblood, indicating the presence of diseased tissue. Definitive diagnosisis generally accomplished by biopsy of the prostate tissue.

No single gene or protein has been identified which is responsible forthe etiology of all prostate cancers. Although PSA is widely used as adiagnostic reagent, it has limitations in its sensitivity and itsability to detect early cancers. It is estimated that approximately 20%to 30% of tumors will be missed when PSA is used alone. It is likelythat diagnostic and prognostic markers for prostate cancer disease willinvolve the identification and use of many different genes and geneproducts to reflect its multifactorial origin.

A continuing goal is to characterize the gene expression patterns of thevarious prostate cancers to genetically differentiate them, providingimportant guidance in preventing and treating cancers. Molecularpictures of cancer, such as the pattern of up-regulated genes identifiedherein, provide an important tool for molecularly dissecting andclassifying cancer, identifying drug targets, providing prognosis andtherapeutic information, etc. For instance, an array of polynucleotidescorresponding to genes differentially regulated in prostate cancer canbe used to screen tissue samples for the existence of cancer, tocategorize the cancer (e.g., by the particular pattern observed), tograde the cancer (e.g., by the number of up-regulated genes and theiramounts of expression), to identify the source of a secondary tumor, toscreen for metastatic cells, etc. These arrays can be used incombination with other markers, e.g., PSA, PMSA (prostate membranespecific antigen), or any of the grading systems used in clinicalmedicine.

As indicated by these studies, cancer is a highly diverse disease.Although all cancers share certain characteristics, the underlying causeand disease progression can differ significantly from patient topatient. So far, over a dozen distinct genes have been identified which,when mutant, result in a cancer. In breast cancer, alone, a handful ofdifferent genes have been isolated which either cause the cancer, orproduce a predisposition to it. As a consequence, disease phenotypes fora particular cancer do not look all the same. In addition to thedifferences in the gene(s) responsible for the cancer, heterogeneityamong individuals, e.g., in age, health, sex, and genetic background,can also influence the disease and its progression. Gene penetrance, inparticular, can vary widely among population members. Recent studieshave shown tremendous diversity in gene expression patterns among cancerpatients. For these and other reasons, one gene/polypeptide target alonecan be insufficient to diagnose or treat a cancer. Even a gene which ishighly differentially-expressed and penetrant in cancer patients may notbe so highly expressed in all patients and at all stages of the cancer.By selecting a set of genes and/or the polypeptides they encode, cancerdiagnostics and therapeutics can be designed which effectively diagnoseand treat a population of diseased individuals, rather than only a smallhandful when single genes are targeted.

Differentially Regulated Prostate Genes

Pc0219

Pc0219 codes for a 891 amino acid polypeptide involved in generegulation (e.g., replication, transcription, and RNA processing). It isup-regulated in prostate cancer. The nucleotide and amino acid sequencesof Pc0219 are shown in SEQ ID NOS 1 and 2. It contains a DEXDc domain atabout amino acid positions 552-703, an AAA domain at about amino acidpositions 570-890, and Viral helicase-1 domain at about amino acidpositions 574-585. The mouse homolog is Slfn1 (NM_(—)011407).Polymorphisms for it are listed in Table 1.

All or part of Pc0219 is located in genomic DNA represented by GenBankID: AC022706, BAC-ID: RP 11-47L3, and Contig ID: NT_(—)010736. Thepresent invention relates to any isolated introns and exons that arepresent in such clone. Such introns and exons can be routinelydetermined.

Nucleic acids of the present invention map to chromosomal band 17q12b.There are a number of different disorders which have been mapped to, orin close proximity to, this chromosome location. These include, e.g.,Abdominal obesity-metabolic syndrome; Van Buchem disease; Familialfrontotemporal dementia (FTD). Nucleic acids of the present inventioncan be used as linkage markers, diagnostic targets, therapeutic targets,for any of the mentioned disorders, as well as any disorders or genesmapping in proximity to it.

Pc0370

Pc0370 codes for a 124 amino acid polypeptide which is up-regulated inprostate cancer. It is expressed in many different tissue types. Thenucleotide and amino acid sequences of Pc0370 are shown in SEQ ID NOS 3and 4. It contains a hydrophobic peptide at about amino acids 1-26, aBCL domain at about amino acids 32-64, and a TOP2c domain at about aminoacids 74-113.

All or part of Pc0370 is located in genomic DNA represented by GenBankID: AC004686/BAC-ID: hRPC.1073_F_(—)15 (for cDNA 122-4270 bp) and ContigID: NT_(—)010740(122-4270 bp). The present invention relates to anyisolated introns and exons that are present in such clone. Such intronsand exons can be routinely determined.

Nucleic acids of the present invention map to chromosomal band 17q22.There are a number of different disorders which have been mapped to, orin close proximity to, this chromosome location. These include, e.g.,Trichodontoosseous syndrome; Gliosis, familial progressive subcortical;Elliptocytosis, Malaysian-Melanesian type; Acanthocytosis, one form;White sponge nevus; Spherocytosis, hereditary; Renal tubular acidosis,distal; Hypertension, essential; Epidermolytic hyperkeratosis; Hcmolyticanemia due to band 3 defect; Pseudohypoaldosteronism type II; Patellaaplasia or hypoplasia; Synostoses syndrome, multiple; Mulibrey nanism;Meckel syndrome; Pituitary tumor, invasive; Placental lactogendeficiency; Isolated growth hormone deficiency, Illig type with absentGH and Kowarski type with bioinactive GH. Nucleic acids of the presentinvention can be used as linkage markers, diagnostic targets,therapeutic targets, for any of the mentioned disorders, as well as anydisorders or genes mapping in proximity to it.

Pc0378

Pc0378 codes for a polypeptide containing 1821 amino which isup-regulated in prostate cancer. The nucleotide and amino acid sequencesof Pc0378 are shown in SEQ ID NOS 5 and 6. It contains eight WD40domains at about amino acid positions: 172-211, 214-253,256-299,310-349,354-393, 408-452,455495, and 498-542; a coiled coil domain atabout amino acid positions 871-908; and tandem bromodomains at aboutamino acid positions 1158-1261 and 1318-1423. It is also expressed innormal adrenal gland, bone marrow, colon, and lymphocytes.

Two cDNAs representing Pc0378 had been previously reported, but neitherhad been recognized as been partial clones. NM_(—)017934 is missing theN-terminal 1114 amino acids and AAG45146 is missing the N-terminal 959amino acids. See, FIG. 1. A partial clone has been reported to interactwith IRS-1. See, e.g., Farhang-Fallah et al., J. Biol. Chem.,275:40492-40497, 2000. The present invention relates to the entirefull-length sequence of PC0378, as well as fragments of it, including,e.g., polynucleotide and DNA-encoding fragments for amino acid positions1-959, 960-1114, 1-1114, 960-1821, 1114-1821, 172-211, 214-253, 256-299,310-349, 354-393, 408-452, 455-495, 498-542, 871-908, etc. Additionalfragments include those comprising polymorphic differences betweenPc078, NM_(—)017934, and AAG45146, e.g., 1150-1180, 1280-1290,1460-1490, 1490-1510, 1590-1620, etc. See, FIG. 1.

All or part of Pc0378 is located in genomic DNA represented by GenBankID: AL356776, BAC-ID: RP11-1477E3, and Contig ID: NT_(—)023399. Thepresent invention relates to any isolated introns and exons that arepresent in such clone which can be routinely determined. Its chromosomalmap position is 6q14.1-q14.3 with a physical position of UniSTS: 80944at 5′ at 84.870 Mb amd UniSTS: 91580 at 3′ at 84.601 Mb The chromosomallocation of Pc078 places it in proximity to several disease loci, e.g.,chorioretinal atrophy, Leber congeneital amaurosis, rod dystrophy 7,macular dystrophy/degeneration, schizophrenia, and cardiomyopathy.Nucleic acids of the present invention can be used as linkage markers,diagnostic targets, therapeutic targets, for any of the mentioneddisorders, as well as any disorders or genes mapping in proximity to it.

Pc444

Pc444 codes for a 235 amino acid polypeptide. The nucleotide and aminoacid sequences of Pc444 are shown in SEQ ID NOS 7 and 8. Polymorphismsfor it are listed in Table 1.

All or part of Pc444 is located in genomic DNA represented by GenBankID: AF236876/BAC-ID: CTD-3048P3, and Contig ID: NT_(—)008043. Thepresent invention relates to any isolated introns and exons that arepresent in such clone. Such introns and exons can be routinelydetermined.

Nucleic acids of the present invention map to chromosomal band8p23.2-p23.3. There are a number of different disorders which have beenmapped to, or in close proximity to, this chromosome location. Theseinclude, e.g., Diamond-blackfan anemia 2, Keratolytic winter erythema(2), asthma-susceptibility loci, human hepatocellular carcinoma, andbladder cancer. Nucleic acids of the present invention can be used aslinkage markers, diagnostic targets, therapeutic targets, for any of thementioned disorders, as well as any disorders or genes mapping inproximity to it.

Pc011

Pc011 codes for a 143 amino acid polypeptide that is up-regulated inprostate cancer. It is also expressed in normal prostate, heart, andtestis. It is related, e.g., to XM_(—)046729 and LOC92700. Thenucleotide and amino acid sequences of Pc011 are shown in SEQ ID NOS 9and 10. Polymorphisms for it are listed in Table 1.

All or part of Pc011 is located in genomic DNA represented by GenBankID: AC007563, BAC-ID: RP 1′-506C8, and Contig ID: NT_(—)005337. Thepresent invention relates to any isolated introns and exons that arepresent in such clone. Such introns and exons can be routinelydetermined.

Nucleic acids of the present invention map to chromosomal band 2q22-q23.There are a number of different disorders which have been mapped to, orin close proximity to, this chromosome location. These include, e.g.,Spastic cerebral palsy, symmetric; Nemaline myopathy 2, autosomalrecessive; Hirschsprung disease with microcephaly, mental retardation,and distinct facial features; Epilepsy, juvenile myoclonic; Epilepsy,generalize idiopathic; Ataxia, episodic; Convulsions, familial febrile;Deafness, autosomal dominant. Nucleic acids of the present invention canbe used as linkage markers, diagnostic targets, therapeutic targets, forany of the mentioned disorders, as well as any disorders or genesmapping in proximity to it.

Pc018

Pc018 codes for a 444-amino acid polypeptide which is related toNM_(—)022346 and XM_(—)039166. It is up-regulated in prostate cancer,and is also expressed in bone marrow, thymus, and testis. The nucleotideand amino acid sequences of Pc018 are shown in SEQ ID NOS 11 and 12. Itcontains a coiled coil domain at about amino acid positions 22-55 and aUVR domain at about amino acid positions 5-40.

All or part of Pc018 is located in genomic DNA represented by GenBankID: AC027576, BAC-ID: RP 11-162M 10, and Contig ID: NT_(—)006344. Thepresent invention relates to any isolated introns and exons that arepresent in such clone. Such introns and exons can be routinelydetermined.

Nucleic acids of the present invention map to chromosomal band 4p 15.3.There are a number of different disorders which have been mapped to, orin close proximity to, this chromosome location. These include, e.g.,Parkinson disease and Huntington-like neurodegenerative disorder 2.Nucleic acids of the present invention can be used as linkage markers,diagnostic targets, therapeutic targets, for any of the mentioneddisorders, as well as any disorders or genes mapping in proximity to it.

Pc287

Pc287 codes for a 272 amino acid polypeptide which is up-regulated inprostate cancer. It is related to NM_(—)033255 which is breastepithelial stromal interaction protein. The nucleotide and amino-acidsequences of Pc287 are shown in SEQ ID NOS 13 and 14. It containstransmembrane domains at about amino acid positions 212-231 and 241-263,and tandem Coiled coil domains at about amino acid positions 6-66 and99-138. Polymorphisms for it are listed in Table 1.

All or part of Pc287 is located in genomic DNA represented by GenBankID: AL137878/BAC-ID: RP11-145J3, GenBank ID: AL4452.17/BAC-ID: RP11-335G18, and Contig ID: NT_(—)009935. The present invention relates toany isolated introns and exons that are present in such clone. Suchintrons and exons can be routinely determined. The physical position ofPc287 is marked by UniSTS:169255 at 5′ at 42.25 Mb and UniSTS:186617 at3′ at 42.10 Mb.

Nucleic acids of the present invention map to chromosomal band 13q14.1.There are a number of different disorders which have been mapped to, orin close proximity to, this chromosome location. These include, e.g.,Rhabdomyosarcoma, alveolar; Nonsmall cell lung cancer; Bladder cancer;Osteosarcoma; and Retinoblastoma. Nucleic acids of the present inventioncan be used as linkage markers, diagnostic targets, therapeutic targets,for any of the mentioned disorders, as well as any disorders or genesmapping in proximity to it.

Pc0382

Pc0382 codes for a 1584 amino acid nuclear-regulatory polypeptide whichis up-regulated in prostate cancer. The nucleotide and amino acidsequences of Pc0382 are shown in SEQ ID NOS 15 and 16. It contains a SAMdomain at about amino acid positions 11-78, a Bipartite nuclearlocalization signal domain at about amino acid positions 269-286, and aKinesin domain at about amino acid positions 1079-1103. Polymorphismsfor it are listed in Table 1.

All or part of Pc0382 is located in genomic DNA represented by GenBankID: AC000119/BAC-ID: RG104104, and Contig ID: NT_(—)029333. The presentinvention relates to any isolated introns and exons that are present insuch clone. Such introns and exons can be routinely determined.

Nucleic acids of the present invention map to chromosomal band 7q21-q22.There are a number of different disorders which have been mapped to, orin close proximity to, this chromosome location. These include, e.g.,SHFM 1 and SHFM ID, Malignant hyperthermia susceptibility, Myoclonicdystonia-11, and Zellweger syndrome. Nucleic acids of the presentinvention can be used as linkage markers, diagnostic targets,therapeutic targets, for any of the mentioned disorders, as well as anydisorders or genes mapping in proximity to it.

Pc036-2

Pc036-2 (NM_(—)006924) codes for a pre-mRNA splicing factor containing248 amino acids. The nucleotide and amino acid sequences of Pc036-2 areshown in SEQ ID NOS 17 and 18. It contains a RNA recognition motif atabout nucleotide positions 176-337. A mouse homolog is X66091.

All or part of Pc036-2 is located in genomic DNA represented by GenBankID: AC021455, BAC-ID: RP11-142B17, and Contig ID: NT_(—)010651. Thepresent invention relates to any isolated introns and exons that arepresent in such clone. Such introns and exons can be routinelydetermined. Using UniSTS probes, Pc036-2 can be chromosomally mapped atits 5′ end with UniSTS: 92217 to 58.475 Mb, and at its 3′ end withUniSTS: 29865 to 58.490 Mb. It maps to chromosomal band 17q21.3-q22.

Pc168

Pc168 (related to NM_(—)033255) codes for a Cu/Zn superoxide dismutase(SOD). Its nucleotide and amino acid sequences are shown in SEQ ID NOS19 and 20.

Pc176

Pc176 (related to AK001739) codes for a polypeptide. Its nucleotide andamino acid sequences are shown in SEQ ID NOS 21 and 22.

Pc345

Pc345 (related to AF220047) is a nucleotide sequence. Its nucleotidesequence is shown in SEQ ID NO 23.

Pc380

Pc380 (related to NM_(—)015836.1) codes for a tryptophanyl tRNAsynthase. It is found in the mitochondria, although it is coded for by anuclear gene. The amino acid and nucleotide sequences of Pc380 are shownin SEQ ID NOS 24 and 25.

Pc513

Pc513 (related to NM_(—)001417) codes for a eukaryotic translationfactor 4B (EIF4B) containing containing 608 amino acids. The nucleotideand amino acid sequences of Pc513 are shown in SEQ ID. NOS 26 and 27. Itcontains a coiled coil domain at about amino acid positions 367-394. The3′ UTR is longer than NM_(—)001417 and contains another gene, NM 018507,in the opposite orientation. A UniGene cluster is represented byHs.93379. A mouse homolog is BC007171.

All or part of Pc513 is located in genomic DNA represented by GenBankID: AC073573, BAC-ID: RP11-1136G11, and Contig ID: NT_(—)030118.1. Thepresent invention relates to any isolated introns and exons that arepresent in such clone. Such introns and exons can be routinelydetermined. Using UniSTS probes, Pc513 can be chromosomally mapped atits 5′ end with UniSTS: 72132 to 55.307 Mb, and its 3′ end with UniSTS:182629 to 55.247 Mb.

In addition to its association with prostate cancer, Pc513 expressioncan be affected in other disorders, as well, including other diseases ofprostate. For example, expression of Pc513 can be detected in manydifferent tissues. Thus, this gene has an additional functional role inthese tissues, and can be involved with diseases associated with them,as well. For instance, Pc513 is involved in other cancers, such asglioma and melanoma.

Nucleic acids of the present invention map to chromosomal band 12q13.11-q 14.3. There are a number of different disorders which have beenmapped to, or in close proximity to, this chromosome location. Theseinclude, e.g., Wagner syndrome (type II), Kniest dysplasia, Sticklersyndrome (type I), achondrogenesis-hypochondrogenesis (type II), SMEDStrudwick type, precocious osteoarthrosis, SED congenita, myxoidliposarcoma, glycogen storage disease VII, scapuloperoneal syndrome(myopathic type), fundus albipunctatus, glioma, noctumal enuresis,melanoma, Sanfilippo syndrome (type D), and pseudovitamin D deficiencyrickets 1. Nucleic acids of the present invention can be used as linkagemarkers; diagnostic targets, therapeutic targets, for any of thementioned disorders, as well as any disorders or genes mapping inproximity to it.

Pc520

Pc520 (related to NM_(—)004111) codes for a flap-structure-specificendonuclease containing 380 amino acids. The nucleotide and amino acidsequences of Pc520 are shown in SEQ ID NOS 28 and 29. It contains a XPGNdomain at about amino acid positions 1-107, a Coiled coil at about aminoacid positions 94-140, a XPGI domain at about amino acid positions146-218, a HhH2 domain at about amino acid positions 220-253, and a53EXOc domain at about amino acid positions 29-297. A UniGene cluster isrepresented by Hs.4756. A mouse homolog is BC010203.

All or part of Pc520 is located in genomic DNA represented by GenBankID: AC004770, BAC-ID: CIT-HSP-311e8, and Contig ID: NT_(—)009296. Thepresent invention relates to any isolated introns and exons that arepresent in such clone. Such introns and exons can be routinelydetermined. Using UniSTS probes, Pc520 can be chromosomally mapped atits 5′ end with UniSTS: 11341 to 60.912 Mb, and its 3′ end with UniSTS:57379 to 60.923 Mb.

In addition to its association with prostate cancer, Pc520 expressioncan be affected in other disorders, as well, including other diseases ofprostate. For example, expression of Pc520 can be detected in manydifferent tissues. Thus, this gene has an additional functional role inthese tissues, and can consequently be involved with diseases associatedwith them, as well.

Nucleic acids of the present invention map to chromosomal band 11q12.2a. There are a number of different disorders which have been mappedto, or in close proximity to, this chromosome location. These include,e.g., angioedema, bone mineral density variability 1, xerodermapigmentosum (group E, subtype 2), Smith-Lemli-Opitz syndrome,osteoporosis-pseudoglioma syndrome, and retinitis pigmentosa. Nucleicacids of the present invention can be used as linkage markers,diagnostic targets, therapeutic targets, for any of the mentioneddisorders, as well as any disorders or genes mapping in proximity to it.

Pc118-2

Pc118-2 (related to XM_(—)027365) codes for an ADP-ribosylation-likefactor containing 203 amino acids. The nucleotide and amino acidsequences of Pc 118-2 are shown in SEQ ID NOS 30 and 31. It containshydrophobic transmrnembrane-like domains at about amino acid positions42-61,66-88, 136-153, and 158-180. It is also related to UniGene clusterHs.75249 and a partial sequence disclosed in WO/0140269. A mouse homologis AF223953.

All or part of Pc 18-2 is located in genomic DNA represented by GenBankID: AC025289, BAC-ID: RP 11-533D19, and Contig ID: NT_(—)024822.4. Thepresent invention relates to any isolated introns and exons that arepresent in such clone. Such introns and exons can be routinelydetermined. Using UniSTS probes, Pc118-2 can be chromosomally mapped atits 5′ end with UniSTS: 6682 to 18.297 Mb, and its 3′ end with UniSTS:16634 to −18.300 Mb.

In addition to its association with prostate cancer, Pc 18-2 expressioncan be affected in other disorders, as well, including other diseases ofprostate. For example, expression of Pc 18-2 can be detected in manydifferent tissues. Thus, this gene has an additional functional role inthese tissues, and may be involved with diseases associated with them,as well. See, e.g. WO/0140269 for a suggested role in other cancers.

Nucleic acids of the present invention map to chromosomal band 16p 12-p13.1. There are a number of different disorders which have been mappedto, or in close proximity to, this chromosome location. These include,e.g., Brody myopathy, medullary cystic kidney disease 2,ceroid-lipofuscinosis, hepatic glycogenosis, familial mitral valveprolapse, atopy susceptibility, retinitis pigmentosa-22, convulsions(e.g., infantile and paroxysmal choreoathetosis), familial Mediterraneanfever, MHC class II deficiency complementation group A, osteopetrosis,epilepsyrn and pseudoxanthoma elasticum. Nucleic acids of the presentinvention can be used as linkage markers, diagnostic targets,therapeutic targets, for any of the mentioned disorders, as well as anydisorders or genes mapping in proximity to it.

Pc242

Pc242 codes for a cytochrome-related protein containing 63 amino acids.The nucleotide and amino acid sequences of Pc242 are shown in SEQ ID NOS32 and 33. It comprises a cytochrome C complex domain at about aminoacids 3-63. A UniGene cluster is Hs.284292.

All or part of Pc242 is located in genomic DNA represented by GenBankID: AC004882, BAC-ID: RP1-76B20, and Contig ID: NT_(—)011520. Thepresent invention relates to any isolated introns and exons that arepresent in such clone. Such introns and exons can be routinelydetermined. Using UniSTS probes, Pc242 can be chromosomally mapped atits 5′ end with UniSTS: 88056 to 26.923 Mb, and at its 3′ end withUniSTS: 15837 to 26.962 Mb.

In addition to its association with prostate cancer, Pc242 expression isassociated with other cancers, e.g., neuroepithelioma and Ewing sarcoma.In addition, its expression can be perturbed in other disorders, aswell, including other diseases of prostate. For example, expression ofPc242 can be detected in many different tissues, although moreabundantly in muscle and heart. Thus, this gene has a functional role inthese tissues, and can be involved with diseases associated with them,as well.

Nucleic acids of the present invention map to chromosomal band 22q 12.There are a number of different disorders which have been mapped to, orin close proximity to, this chromosome location. These include, e.g.,macrothrombocytopathy with nephritis and deafness, epilepsy (partial,with variable foci), schizophrenia, Epstein syndrome, Ewing sarcoma,heme oxygenase-1 deficiency, and neuroepithelioma. Nucleic acids of thepresent invention can be used as linkage markers, diagnostic targets,therapeutic targets, for any of the mentioned disorders, as well as anydisorders or genes mapping in proximity to it.

Pc455

Pc455 codes for a polypeptide having 316 amino acids. The nucleotide andamino acid sequences of Pc455 are shown in SEQ ID NOS 34 and 35.

Pc143

Pc143 codes for a polypeptide having 307 amino acids. The nucleotide andamino acid sequences of Pc143 are shown in SEQ ID NOS 36 and 37.

Pc099

Pc099 codes for a teneurin-2, a polypeptide containing 1351 amino acids.It is a transmembrane protein. The nucleotide and amino acid sequencesof Pc099 are shown in SEQ ID NOS 38 and 39. It contains EGF-like domainsat about amino acid positions, 177-205, 208-236, 241-270, 273-302,307-337, and 340-372. A partial human clone is XM 047995 (See, FIG. 2),and a UriGene cluster is Hs. 173560. The mouse homolog is NM_(—)011856and the rat homolog is NM_(—)020088.

All or part of Pc099-2 is located in genomic DNA represented by GenBankID: AC025764, BAC-ID: CTB-41A10, and Contig ID: NT_(—)006907. Thepresent invention relates to any isolated introns and exons that arepresent in such clone. Such introns and exons can be routinelydetermined. Using UniSTS probes, Pc099-2 can be chromosomally mapped atits 5′ end with UniSTS: 37515 to 170.680 Mb, and at its 3′ end withUniSTS: 9147 to 170.865 Mb.

In addition to its association with prostate cancer, Pc099-2 expressioncan be affected in other disorders, as well, including other diseases ofprostate. For example, expression of Pc099-2 can be detected in brainand heart. Thus, this gene has a functional role in these tissues, andmay be involved with diseases associated with them, as well. Forinstance, it can have a role in making and/or maintaining neuronalconnections, and in forming developing tissues.

Nucleic acids of the present invention map to chromosomal band 5q34.There are a number of different disorders which have been mapped to, orin close proximity to, this chromosome location. These include, e.g.,obesity susceptibility to, nocturnal asthma susceptibility, limb-girdletype 2F muscular dystrophy, carnitine deficiency, predisposition tomyeloid malignancy, atrial septal defect with atrioventricularconduction defects, parietal foramina 1, and type 2 craniosynostosis.Nucleic acids of the present invention can be used as linkage markers,diagnostic targets, therapeutic targets, for any of the mentioneddisorders, as well as any disorders or genes mapping in proximity to it.

Pc452

Pc452 codes for a polypeptide containing 745 amino acid. Its nucleotideand amino acid sequences are shown in SEQ ID NOS 40 and 41. It plays arole in nuclear regulation, and therefore the polypeptide it encodes canbe identified in the cell's nucleus. Partial sequences for it arerepresented by UniGene Hs.99807. A mouse homolog is partial cloneAK012040.

All or part of Pc452 is located in genomic DNA represented by GenBankID: AC067802, BAC-ID: RP11-1434113, and Contig ID: NT_(—)019306.6. UsingUniSTS probes, Pc452 can be chromosomally mapped at its 5′ end withUniSTS: 27526 to 109.366 Mb, and at its 3′ end with UniSTS: 50791 to109.33 Mb. The present invention relates to any isolated introns andexons that are present in such clone. Such introns and exons can beroutinely determined Disorders associated with Pc452 can affectprostate, as well as other tissues and cell types in the body. Such geneeffects can be caused by the direct action of the gene on another tissueor cell type, or indirectly, e.g., where a prostate tissue dysfunctionor abnormality has downstream effects on other systems and cell types inthe body. Furthermore, levels of Pc452 expression can occur in celltypes other than prostate, and thus can have a function outside of it.For instance, expression of Pc452 can be detected in bone marrow,testis, and thymus.

Nucleic acids of the present invention map to chromosomal band 2q 13-q14.1. There are a number of different disorders which have been mappedto, or in close proximity to, this chromosome location. These include,e.g., congenital hypothyroidism due to thyroid dysgenesis or hypoplasia,nephronophthisis, neonatal purpura fulminans, thrombophilia due toprotein C deficiency, distal hereditary motor neuronopathy, colorectalcancer with chromosomal instability, retinitis pigmentosa(MERTK-related), hepatocellular carcinoma, and dilated cardiomyopathy.Nucleic acids of the present invention can be used as linkage markers,diagnostic targets, therapeutic targets, for any of the mentioneddisorders, as well as any disorders or genes mapping in proximity to it.

Pc066

Pc066 (SEQ ID NO 42 and 43; related to NM_(—)019896), codes for a DNApolymerase epsilon p12 subunit. It is up-regulated in prostate cancer.

Pc0393

Pc0393 (SEQ ID NO 44 and 45; related to XM_(—)064903) is up-regulated inprostate cancer.

Pc204

Pc240 (SEQ ID NO 46 and 47; related to U10117) codes forendothelial-monocyte activating polypeptide 11 (small inducible cytokinesubfamily E, member 1) and is down-regulated in prostate cancer.

Pc432

Pc432 (SEQ ID NO 48 and 49; related to X07109) codes for a proteinkinase C (PKC) type beta II, and is down-regulated in prostate cancer.

Nucleic Acids

In accordance with the present invention, genes have been identifiedwhich are differentially expressed in prostate cancer. These genes canbe further divided into groups based on additional characteristics oftheir expression and the tissues in which they are expressed. Forinstance, genes can be further subdivided based on the stage and/orgrade of the cancer in which they are expressed. Genes can also begrouped based on their penetrance in a prostate cancer, e.g., expressedin all prostate cancer examined, expressed in a certain percentage ofprostate cancer examined, etc. These groupings do not restrict or limitthe use such genes in therapeutic, diagnostic, prognostic, etc.,applications. For instance, a gene which is expressed in only somecancers (e.g., incompletely penetrant) may be useful in therapeuticapplications to treat a subset of cancers. Similarly, a co-penetrantgene, or a gene which is expressed in prostate cancer and other normaltissues, may be useful as a therapeutic or diagnostic, even if itsexpression pattern is not highly prostate specific. Thus, the uses ofthe genes or their products are not limited by their patterns ofexpression.

By the phrase “differential expression,” it is meant that the levels ofexpression of a gene, as measured by its transcription or translationproduct, are different depending upon the specific cell-type or tissue(e.g., in an averaging assay that looks at a population of cells). Thereare no absolute amounts by which the gene expression levels must vary,as long as the differences are measurable.

The phrase “up-regulated” indicates that an mRNA transcript or othernucleic acid corresponding to a polynucleotide of the present inventionis expressed in larger amounts in a cancer as compared to the sametranscript expressed in normal cells from which the cancer was derived.In general, up-regulation can be assessed by any suitable method,including any of the nucleic acid detection and hybridization methodsmentioned below, as well as polypeptide-based methods. Up-regulationalso includes going from substantially no expression in a normal tissue,from detectable expression in a normal tissue, from significantexpression in a normal tissue, to higher levels in the cancer.

Differential regulation can be determined by any suitable method, e.g.,by comparing its abundance per gram of RNA (e.g., total RNA,polyadenylated mRNA, etc.) extracted from a prostate tissue incomparison to the corresponding normal tissue. The normal tissue can befrom the same or different individual or source. For convenience, it canbe supplied as a separate component or in a kit in combination withprobes and other reagents for detecting genes. The quantity by which anucleic acid is differentially-regulated can be any value, e.g., about10% more or less of normal expression, about 50% more or less of normalexpression, 2-fold more or less, 5-fold more or less, 10-fold more orless, etc.

The amount of transcript can also be compared to a different gene in thesame sample, especially a gene whose abundance is known andsubstantially no different in its expression between normal and cancercells (e.g., a “control” gene). If represented as a ratio, with thequantity of differentially-regulated gene transcript in the numeratorand the control gene transcript in the denominator, the ratio would belarger, e.g., in prostate cancer than in a sample from normal prostatetissue.

Differential-regulation can arise through a number of differentmechanisms. The present invention is not bound by any specific waythrough which it occurs. Differential-regulation of a polynucleotide canoccur, e.g., by modulating (1) transcriptional rate of the gene (e.g.,increasing its rate, inducing or stimulating its transcription from abasal, low-level rate, etc.), (2) the post-transcriptional processing ofRNA transcripts, (3) the transport of RNA from the nucleus into thecytoplasm, (4) the RNA nuclear and cytoplasmic turnover (e.g., by virtueof having higher stability or resistance to degradation), andcombinations thereof. See, e.g., Tollervey and Caceras, Cell,103:703-709, 2000.

A differentially-regulated polynucleotide is useful in a variety ofdifferent applications as described in greater details below. Because itis more abundant in cancer, it and its expression products can be usedin a diagnostic test to assay for the presence of cancer, e.g., intissue sections, in a biopsy sample, in total RNA, in lymph, in blood,etc. Differentially-regulated polynucleotides and polypeptides can beused individually, or in groups, to assess the cancer, e.g., todetermine the specific type of cancer, its stage of development, thenature of the genetic defect, etc., or to assess the efficacy of atreatment modality. How to use polynucleotides in diagnostic andprognostic assays is discussed below. In addition, the polynucleotidesand the polypeptides they encode, can serve as a target for therapy ordrug discovery. A polypeptide, coded for by a differentially-regulatedpolynucleotide, which is displayed on the cell-surface, can be a targetfor immunotherapy to destroy, inhibit, etc., the diseased tissue.Differentially-regulated transcripts can also be used in drug discoveryschemes to identify pharmacological agents which suppress, inhibit,etc., their differential-regulation, thereby preventing the phenotypeassociated with their expression. Thus, a differentially-regulatedpolynucleotide and its expression products of the present invention havesignificant applications in diagnostic, therapeutic, prognostic, drugdevelopment, and related areas.

The expression patterns of the differentially expressed genes disclosedherein can be described as a “fingerprint” in that they are adistinctive pattern displayed by a cancer. Just as with a fingerprint,an expression pattern can be used as a unique identifier to characterizethe status of a tissue sample. The list of genes represented by SEQ IDNO 1-49 provides an example of a cell expression profile for a prostatecancer. It can be used as a point of reference to compare andcharacterize unknown samples and samples for which further informationis sought. Tissue fingerprints can be used in many ways, e.g., toclassify an unknown tissue as being a prostate cancer, to determine theorigin of a particular cancer (e.g., the origin of metastatic cells), todetermine the presence of a cancer in a biopsy sample, to assess theefficacy of a cancer therapy in a human patient or a non-human animalmodel, to detect circulating cancer cells in blood or a lymph nodebiopsy, etc. While the expression profile of the complete gene setrepresented by SEQ ID NO 1-49 may be most informative, a fingerprintcontaining expression information from less than the full collection canbe useful, as well. In the same way that an incomplete fingerprint maycontain enough of the pattern of whorls, arches, loops, and ridges, toidentify the individual, a cell expression fingerprint containing lessthan the full complement may be adequate to provide useful and uniqueidentifying and other information about the sample. Moreover, cancer isa multifactorial disease, involving genetic aberrations in more thangene locus. This multifaceted nature may be reflected in different cellexpression profiles associated with prostate cancers arising indifferent individuals, in different locations in the same individual, oreven within the same cancer locus. As a result, a complete match with aparticular cell expression profile, as shown herein, is not necessary toclassify a cancer as being of the same type or stage. Similarity to onecell expression profile, e.g., as compared to another, can be adequateto classify cancer types, grades, and stages.

A mammalian polynucleotide, or fragment thereof, of the presentinvention is a polynucleotide having a nucleotide sequence obtainablefrom a natural source. It therefore includes naturally-occurring normal,naturally-occurring mutant, and naturally-occurring polymorphic alleles(e.g., SNPs), differentially-spliced transcripts, splice-variants, etc.By the term “naturally-occurring,” it is meant that the polynucleotideis obtainable from a natural source, e.g., animal tissue and cells, bodyfluids, tissue culture cells, forensic samples. Natural sources include,e.g., living cells obtained from tissues and whole organisms, tumors,cultured cell lines, including primary and immortalized cell lines.Naturally-occurring mutations can include deletions (e.g., a truncatedamino- or carboxy-terminus), substitutions, inversions, or additions ofnucleotide sequence. These genes can be detected and isolated bypolynucleotide hybridization according to methods which one skilled inthe art would know, e.g., as discussed below.

A polynucleotide according to the present invention can be obtained froma variety of different sources. It can be obtained from DNA or RNA, suchas polyadenylated mRNA or total RNA, e.g., isolated from tissues, cells,or whole organism. The polynucleotide can be obtained directly from DNAor RNA, from a cDNA library, from a genomic library, etc. Thepolynucleotide can be obtained from a cell or tissue (e.g., from anembryonic or adult tissues) at a particular stage of development, havinga desired genotype, phenotype, disease status, etc.

The polynucleotides described in SEQ ID NO 1, 3, 5, 7, 9, 11, 13, 15,17, 19, 21, 23, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 44, 46, or 48can be partial sequences that correspond to full-length,naturally-occurring transcripts. The present invention includes, aswell, full-length polynucleotides that comprise these partial sequences,e.g., genomic DNAs and polynucleotides comprising a start and stopcodon, a start codon and a polyA tail, a transcription start and a polyAtail, etc. These sequences can be obtained by any suitable method, e.g.,using a partial sequence as a probe to select a fill-length cDNA from alibrary containing full-length inserts. A polynucleotide which “codeswithout interruption” refers to a polynucleotide having a continuousopen reading frame (“ORF”) as compared to an ORF which is interrupted byintrons or other noncoding sequences.

Polynucleotides and polypeptides can be excluded as compositions fromthe present invention if, e.g., listed in a publicly available databaseson the day this application was filed and/or disclosed in a patentapplication having an earlier filing or priority date than thisapplication and/or conceived and/or reduced to practice earlier than apolynucleotide in this application.

As described herein, the phrase “an isolated polynucleotide which is SEQID NO,” or “an isolated polynucleotide which is selected from SEQ IDNO,” refers to an isolated nucleic acid molecule from which the recitedsequence was derived (e.g., a cDNA derived from mRNA; cDNA derived fromgenomic DNA). Because of sequencing errors, typographical errors, etc.,the actual naturally-occurring sequence may differ from a SEQ ID listedherein. Thus, the phrase indicates the specific molecule from which thesequence was derived, rather than a molecule having that exact recitednucleotide sequence, analogously to how a culture depository numberrefers to a specific cloned fragment in a cryotube.

As explained in more detail below, a polynucleotide sequence of theinvention can contain the complete sequence as shown in SEQ ID NO 1, 3,5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 24, 26, 28, 30, 32, 34, 36, 38, 40,42, 44, 46, or 48, degenerate sequences thereof, anti-sense, muteinsthereof, genes comprising said sequences, full-length cDNAs comprisingsaid sequences, complete genomic sequences, fragments thereof, homologs,primers, nucleic acid molecules which hybridize thereto, derivativesthereof, etc.

Genomic

The present invention also relates genomic DNA from which thepolynucleotides of the present invention can be derived. A genomic DNAcoding for a human, mouse, or other mammalian polynucleotide, can beobtained routinely, for example, by screening a genomic library (e.g., aYAC library) with a polynucleotide of the present invention, or bysearching nucleotide databases, such as GenBank and EMBL, for matches.Promoter and other regulatory regions (including both 5′ and 3′sequences) can be identified upstream or down stream of coding andexpressed RNAs, and assayed routinely for activity, e.g., by joining toa reporter gene (e.g., CAT, GFP, alkaline phosphatase, luciferase,galatosidase). A promoter obtained from a prostate selective gene can beused, e.g., in gene therapy to obtain tissue-specific expression of aheterologous gene (e.g., coding for a therapeutic product or cytotoxin).5′ and 3′ sequences (including, UTRs and introns) can be used tomodulate or regulate stability, transcription, and translation ofnucleic acids, including the sequence to which is attached in nature, aswell as heterologous nucleic acids.

Constructs

A polynucleotide of the present invention can comprise additionalpolynucleotide sequences, e.g., sequences to enhance expression,detection, uptake, cataloging, tagging, etc. A polynucleotide caninclude only coding sequence; a coding sequence and additionalnon-naturally occurring or heterologous coding sequence (e.g., sequencescoding for leader, signal, secretory, targeting, enzymatic, fluorescent,antibiotic resistance, and other functional or diagnostic peptides);coding sequences and non-coding sequences, e.g., untranslated sequencesat either a 5′ or 3′ end, or dispersed in the coding sequence, e.g.,introns.

A polynucleotide according to the present invention also can comprise anexpression control sequence operably linked to a polynucleotide asdescribed above. The phrase “expression control sequence” means apolynucleotide sequence that regulates expression of a polypeptide codedfor by a polynucleotide to which it is functionally (“operably”) linked.Expression can be regulated at the level of the mRNA or polypeptide.Thus, the expression control sequence includes mRNA-related elements andprotein-related elements. Such elements include promoters, enhancers(viral or cellular), ribosome binding sequences, transcriptionalterminators, etc. An expression control sequence is operably linked to anucleotide coding sequence when the expression control sequence ispositioned in such a manner to effect or achieve expression of thecoding sequence. For example, when a promoter is operably linked 5′ to acoding sequence, expression of the coding sequence is driven by thepromoter. Expression control sequences can include an initiation codonand additional nucleotides to place a partial nucleotide sequence of thepresent invention in-frame in order to produce a polypeptide (e.g., pETvectors from Promega have been designed to permit a molecule to beinserted into all three reading frames to identify the one that resultsin polypeptide expression). Expression control sequences can beheterologous or endogenous to the normal gene.

A polynucleotide of the present invention can also comprise nucleic acidvector sequences, e.g., for cloning, expression, amplification,selection, etc. Any effective vector can be used. A vector is, e.g., apolynucleotide molecule which can replicate autonomously in a host cell,e.g., containing an origin of replication. Vectors can be useful toperform manipulations, to propagate, and/or obtain large quantities ofthe recombinant molecule in a desired host. A skilled worker can selecta vector depending on the purpose desired, e.g., to propagate therecombinant molecule in bacteria, yeast, insect, or mammalian cells. Thefollowing vectors are provided by way of example. Bacterial: pQE70,pQE60, pQE-9 (Qiagen), pBS, pD10, Phagescript, phiX 174, pBK Phagemid,pNH8A, pNH 16a, pNH 18Z, pNH46A (Stratagene); Bluescript KS+II(Stratagene); ptrc99a, pKK223-3, pKK233-3, pDR540, pRIT5 (Pharmacia).Eukaryotic: PWLNEO, pSV2CAT, pOG44, pXT1, pSG (Stratagene), pSVK3, PBPV,PMSG, pSVL (Pharmacia), pCR2.1/TOPO, pCR11/TOPO, pCR4/TOPO, pTrcHisB,pCMV6-XL4, etc. However, any other vector, e.g., plasmids, viruses, orparts thereof, may be used as long as they are replicable and viable inthe desired host. The vector can also comprise sequences which enable itto replicate in the host whose genome is to be modified.

Hybridization

Polynucleotide hybridization, as discussed in more detail below, isuseful in a variety of applications, including, in gene detectionmethods, for identifying mutations, for making mutations, to identifyhomologs in the same and different species, to identify related membersof the same gene family, in diagnostic and prognostic assays, intherapeutic applications (e.g., where an antisense polynucleotide isused to inhibit expression), etc.

The ability of two single-stranded polynucleotide preparations tohybridize together is a measure of their nucleotide sequencecomplementarity, e.g., base-pairing between nucleotides, such as A-T,G-C, etc. The invention thus also relates to polynucleotides, and theircomplements, which hybridize to a polynucleotide comprising a nucleotidesequence as set forth in SEQ ID NO 1, 3, 5, 7, 9, 11, 13, 15, 17, 19,21, 23, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 44, 46, or 48 andgenomic sequences thereof. A nucleotide sequence hybridizing to thelatter sequence will have a complementary polynucleotide strand, or actas a template for one in the presence of a polymerase (i.e., anappropriate polynucleotide synthesizing enzyme). The present inventionincludes both strands of polynucleotide, e.g., a sense strand and ananti-sense strand.

Hybridization conditions can be chosen to select polynucleotides whichhave a desired amount of nucleotide complementarity with the nucleotidesequences set forth in SEQ ID NO 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21,23, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 44, 46; or 48 and genomicsequences thereof. A polynucleotide capable of hybridizing to suchsequence, preferably, possesses, e.g., about 70%, 75%, 80%, 85%, 87%,90%, 92%, 95%, 97%, 99%, or 100% complementarity, between the sequences.The present invention particularly relates to polynucleotide sequenceswhich hybridize to the nucleotide sequences set forth in SEQ ID NO 1, 3,5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 24, 26, 28, 30, 32, 34, 36, 38, 40,42, 44, 46, or 48 or genomic sequences thereof, under low or highstringency conditions. These conditions can be used, e.g., to selectcorresponding homologs in non-human species.

Polynucleotides which hybridize to polynucleotides of the presentinvention can be selected in various ways. Filter-type blots (i.e.,matrices containing polynucleotide, such as nitrocellulose), glasschips, and other matrices and substrates comprising polynucleotides(short or long) of interest, can be incubated in a prehybridizationsolution (e.g., 6×SSC, 0.5% SDS, 100 μg/ml denatured salmon sperm DNA,5× Denhardt's solution, and 50% formamide), at 22-68° C., overnight, andthen hybridized with a detectable polynucleotide probe under conditionsappropriate to achieve the desired stringency. In general, when highhomology or sequence identity is desired, a high temperature can be used(e.g., 65° C.). As the homology drops, lower washing temperatures areused. For salt concentrations, the lower the salt concentration, thehigher the stringency. The length of the probe is another consideration.Very short probes (e.g., less than 100 base pairs) are washed at lowertemperatures, even if the homology is high. With short probes, formamidecan be omitted. See, e.g., Current Protocols in Molecular Biology,Chapter 6, Screening of Recombinant Libraries; Sambrook et al.,Molecular Cloning, 1989, Chapter 9.

For instance, high stringency conditions can be achieved by incubatingthe blot overnight (e.g., at least 12 hours) with a long polynucleotideprobe in a hybridization solution containing, e.g., about 5×SSC, 0.5%SDS, 100 μg/ml denatured salmon sperm DNA and 50% formamide, at 42° C.Blots can be washed at high stringency conditions that allow, e.g., forless than 5% bp mismatch (e.g., wash twice in 0.1% SSC and 0.1% SDS for30 min at 65° C.), i.e., selecting sequences having 95% or greatersequence identity.

Other non-limiting examples of high stringency conditions includes afinal wash at 65° C. in aqueous buffer containing 30 mM NaCl and 0.5%SDS. Another example of high stringent conditions is hybridization in 7%SDS, 0.5 M NaPO₄, pH 7, 1 mM EDTA at 50° C., e.g., overnight, followedby one or more washes with a 1% SDS solution at 42° C. Whereas highstringency washes can allow for less than 5% mismatch, reduced or lowstringency conditions can permit up to 20% nucleotide mismatch.Hybridization at low stringency can be accomplished as above, but usinglower formamide conditions, lower temperatures and/or lower saltconcentrations, as well as longer periods of incubation time.

Hybridization can also be based on a calculation of melting temperature(Tm) of the hybrid formed between the probe and its target, as describedin Sambrook et al. Generally, the temperature Tm at which a shortoligonucleotide (containing 18 nucleotides or fewer) will melt from itstarget sequence is given by the following equation: Tm=(number of A'sand T's)×2° C.+(number of C's and G's)×4° C. For longer molecules,Tm=81.5+16.6 log₁₀[Na⁺]+0.41 (% GC)-600/N where [Na⁺] is the molarconcentration of sodium ions, % GC is the percentage of GC base pairs inthe probe, and N is the length. Hybridization can be carried out atseveral degrees below this temperature to ensure that the probe andtarget can hybridize. Mismatches can be allowed for by lowering thetemperature even further.

Stringent conditions can be selected to isolate sequences, and theircomplements, which have, e.g., at least about 90%, 95%, or 97%,nucleotide complementarity between the probe (e.g., a shortpolynucleotide of SEQ ID NO 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23,24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 44, 46, or 48 or genomicsequences thereof) and a target polynucleotide.

Other homologs of polynucleotides of the present invention can beobtained from mammalian and non-mammalian sources according to variousmethods. For example, hybridization with a polynucleotide can beemployed to select homologs, e.g., as described in Sambrook et al.,Molecular Cloning, Chapter 11, 1989. Such homologs can have varyingamounts of nucleotide and amino acid sequence identity and similarity tosuch polynucleotides of the present invention. Mammalian organismsinclude, e.g., mice, rats, monkeys, pigs, cows, etc. Non-mammalianorganisms include, e.g., vertebrates, invertebrates, zebra fish,chicken, Drosophila, C. elegans, Xenopus, yeast such as S. pombe, S.cerevisiae, roundworms, prokaryotes, plants, Arabidopsis, artemia,viruses, etc. The degree of nucleotide sequence identity between humanand mouse can be about, e.g. 70% or more, 85% or more for open readingframes, etc.

Alignment

Alignments can be accomplished by using any effective algorithm. Forpairwise alignments of DNA sequences, the methods described byWilbur-Lipman (e.g., Wilbur and Lipman, Proc. Natl. Acad. Sci.,80:726-730, 1983) or Martinez/Needleman-Wunsch (e.g., Martinez, NucleicAcid Res., 11:46294634, 1983) can be used. For instance, if theMartinez/Needleman-Wunsch DNA alignment is applied, the minimum matchcan be set at 9, gap penalty at 1.10, and gap length penalty at 0.33.The results can be calculated as a similarity index, equal to the sum ofthe matching residues divided by the sum of all residues and gapcharacters, and then multiplied by 100 to express as a percent.Similarity index for related genes at the nucleotide level in accordancewith the present invention can be greater than 70%, 80%, 85%, 90%, 95%,99%, or more. Pairs of protein sequences can be aligned by theLipman-Pearson method (e.g., Lipman and Pearson, Science, 227:1435-1441,1985) with k-tuple set at 2, gap penalty set at 4, and gap lengthpenalty set at 12. Results can be expressed as percent similarity index,where related genes at the amino acid level in accordance with thepresent invention can be greater than 65%, 70%, 75%, 80%, 85%, 90%, 95%,99%, or more. Various commercial and free sources of alignment programsare available, e.g., MegAlign by DNA Star, BLAST (National Center forBiotechnology Information), BCM (Baylor College of Medicine) Launcher,etc. BLAST can be used to calculate amino acid sequence identity, aminoacid sequence homology, and nucleotide sequence identity. Thesecalculations can be made along the entire length of each of the targetsequences which are to be compared.

After two sequences have been aligned, a “percent sequence identity” canbe determined. For these purposes, it is convenient to refer to aReference Sequence and a Compared Sequence, where the Compared Sequenceis compared to the Reference Sequence. Percent sequence identity can bedetermined according to the following formula: Percent Identity=100[1-(C/R)], wherein C is the number of differences between the ReferenceSequence and the Compared Sequence over the length of alignment betweenthe Reference Sequence and the Compared Sequence where (i) each base oramino acid in the Reference Sequence that does not have a correspondingaligned base or amino acid in the Compared Sequence, (ii) each gap inthe Reference Sequence, (iii) each aligned base or amino acid in theReference Sequence that is different from an aligned base or amino acidin the Compared Sequence, constitutes a difference; and R is the numberof bases or amino acids in the Reference Sequence over the length of thealignment with the Compared Sequence with any gap created in theReference Sequence also being counted as a base or amino acid.

Percent sequence identity can also be determined by other conventionalmethods, e.g., as described in Altschul et al., Bull. Math. Bio. 48:603-616, 1986 and Henikoff and Henikoff, Proc. Natl. Acad. Sci. USA89:10915-10919, 1992.

Specific Polynucleotide Probes

A polynucleotide of the present invention can comprise any continuousnucleotide sequence of SEQ ID NO 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21,23, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 44, 46, or 48, or sequenceswhich share sequence identity thereto, or complements thereof. The tern“probe” refers to any substance that can be used to detect, identify,isolate, etc., another substance. A polynucleotide probe is comprised ofnucleic acid can be used to detect, identify, etc., other nucleic acids,such as DNA and RNA.

These polynucleotides can be of any desired size that is effective toachieve the specificity desired. For example, a probe can be from about7 or 8 nucleotides to several thousand nucleotides, depending upon itsuse and purpose. For instance, a probe used as a primer PCR can beshorter than a probe used in an ordered array of polynucleotide probes.Probe sizes vary, and the invention is not limited in any way by theirsize, e.g., probes can be from about 7-2000 nucleotides, 7-1000, 8-700,8-600, 8-500, 8-400, 8-300, 8-150, 8-100, 8-75,7-50, 10-25, 14-16, atleast about 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22,23, 24, 25, 25, 26, etc., or more. The polynucleotides can havenon-naturally-occurring nucleotides, e.g., inosine, AZT, 3TC, etc. Thepolynucleotides can have 100% sequence identity or complementarity to asequence of SEQ ID NO 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 24, 26,28, 30, 32, 34, 36, 38, 40, 42, 44, 46, or 48, or it can have mismatchesor nucleotide substitutions, e.g., 1, 2, 3, 4, or 5 substitutions. Theprobes can be single-stranded or double-stranded.

In accordance with the present invention, a polynucleotide can bepresent in a kit, where the kit includes, e.g., one or morepolynucleotides, a desired buffer (e.g., phosphate, tris, etc.),detection compositions, RNA or cDNA from different tissues to be used ascontrols, libraries, etc. The polynucleotide can be labeled orunlabeled, with radioactive or non-radioactive labels as known in theart. Kits can comprise one or more pairs of polynucleotides foramplifying nucleic acids specific for differentially-regulated genes ofthe present invention, e.g., comprising a forward and reverse primereffective in PCR. These include both sense and anti-sense orientations.For instance, in PCR-based methods (such as RT-PCR), a pair of primersare typically used, one having a sense sequence and the other having anantisense sequence.

Another aspect of the present invention is a nucleotide sequence that isspecific to, or for, a selective polynucleotide. The phrases “specificfor” or “specific to” a polynucleotide have a functional meaning thatthe polynucleotide can be used to identify the presence of one or moretarget genes in a sample. It is specific in the sense that it can beused to detect polynucleotides above background noise (“non-specificbinding”). A specific sequence is a defined order of nucleotides (oramino acid sequences, if it is a polypeptide sequence) which occurs inthe polynucleotide, e.g., in the nucleotide sequences of 1, 3, 5, 7, 9,11, 13, 15, 17, 19, 21, 23, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 44,46, or 48, and which is characteristic of that target sequence, andsubstantially no non-target sequences. A probe or mixture of probes cancomprise a sequence or sequences that are specific to a plurality oftarget sequences, e.g., where the sequence is a consensus sequence, afunctional domain, etc., e.g., capable of recognizing a family ofrelated genes. Such sequences can be used as probes in any of themethods described herein or incorporated by reference. Both sense andantisense nucleotide sequences are included. A specific polynucleotideaccording to the present invention can be determined routinely.

A polynucleotide comprising a specific sequence can be used as ahybridization probe to identify the presence of, e.g., human or mousepolynucleotide, in a sample comprising a mixture of polynucleotides,e.g., on a Northern blot. Hybridization can be performed under highstringent conditions (see, above) to select polynucleotides (and theircomplements which can contain the coding sequence) having at least 90%,95%, 99%, etc., identity (i.e., complementarity) to the probe, but lessstringent conditions can also be used. A specific polynucleotidesequence can also be fused in-frame, at either its 5′ or 3′ end, tovarious nucleotide sequences as mentioned throughout the patent,including coding sequences for enzymes, detectable markers, GFP, etc,expression control sequences, etc.

A polynucleotide probe, especially one that is specific to apolynucleotide of the present invention, can be used in gene detectionand hybridization methods as already described. In one embodiment, aspecific polynucleotide probe can be used to detect whether a particulartissue or cell-type is present in a target sample. To carry out such amethod, a selective polynucleotide can be chosen which is characteristicof the desired target tissue. Such polynucleotide is preferably chosenso that it is expressed or displayed in the target tissue, but not inother tissues which are present in the sample. For instance, ifdetection of prostate is desired, it may not matter whether theselective polynucleotide is expressed in other tissues, as long as it isnot expressed in cells normally present in blood, e.g., peripheral bloodmononuclear cells. Starting from the selective polynucleotide, aspecific polynucleotide probe can be designed which hybridizes (ifhybridization is the basis of the assay) under the hybridizationconditions to the selective polynucleotide, whereby the presence of theselective polynucleotide can be determined.

Probes which are specific for polynucleotides of the present inventioncan also be prepared using involve transcription-based systems, e.g.,incorporating an RNA polymerase promoter into a selective polynucleotideof the present invention, and then transcribing anti-sense RNA using thepolynucleotide as a template. See, e.g., U.S. Pat. No. 5,545,522.

Polynucleotide Composition

A polynucleotide according to the present invention can comprise, e.g.,DNA, RNA, synthetic polynucleotide, peptide polynucleotide, modifiednucleotides, dsDNA, ssDNA, ssRNA, dsRNA, and mixtures thereof. Apolynucleotide can be single- or double-stranded, triplex, DNA:RNA,duplexes, comprise hairpins, and other secondary structures, etc.Nucleotides comprising a polynucleotide can be joined via various knownlinkages, e.g., ester, sulfamate, sulfamide, phosphorothioate,phosphoramidate, methylphosphonate, carbamate, etc., depending on thedesired purpose, e.g., resistance to nucleases, such as RNAse H,improved in vivo stability, etc. See, e.g., U.S. Pat. No. 5,378,825. Anydesired nucleotide or nucleotide analog can be incorporated, e.g.,6-mercaptoguanine, 8-oxo-guanine, etc.

Various modifications can be made to the polynucleotides, such asattaching detectable markers (avidin, biotin, radioactive elements,fluorescent tags and dyes, energy transfer labels, energy-emittinglabels, binding partners, etc.) or moieties which improve hybridization,detection, and/or stability. The polynucleotides can also be attached tosolid supports, e.g., nitrocellulose, magnetic or paramagneticmicrospheres (e.g., as described in U.S. Pat. No. 5,411,863; U.S. Pat.No. 5,543,289; for instance, comprising ferromagnetic, supermagnetic,paramagnetic, superparamagnetic, iron oxide and polysaccharide), nylon,agarose, diazotized cellulose, latex solid microspheres,polyacrylamides, etc., according to a desired method. See, e.g., U.S.Pat. Nos. 5,470,967, 5,476,925, and 5,478,893.

Polynucleotide according to the present invention can be labeledaccording to any desired method. The polynucleotide can be labeled usingradioactive tracers such as ³²P, ³⁵S, ³H, or ¹⁴C, to mention somecommonly used tracers. The radioactive labeling can be carried outaccording to any method, such as, for example, terminal labeling at the3′ or 5′ end using a radiolabeled nucleotide, polynucleotide kinase(with or without dephosphorylation with a phosphatase) or a ligase(depending on the end to be labeled). A non-radioactive labeling canalso be used, combining a polynucleotide of the present invention withresidues having immunological properties (antigens, haptens), a specificaffinity for certain reagents (ligands), properties enabling detectableenzyme reactions to be completed (enzymes or coenzymes, enzymesubstrates, or other substances involved in an enzymatic reaction), orcharacteristic physical properties, such as fluorescence or the emissionor absorption of light at a desired wavelength, etc.

Nucleic Acid Detection Methods

Another aspect of the present invention relates to methods and processesfor detecting differentially-regulated genes of the present invention.Detection methods have a variety of applications, including fordiagnostic, prognostic, forensic, and research applications. Toaccomplish gene detection, a polynucleotide in accordance with thepresent invention can be used as a “probe.” The term “probe” or“polynucleotide probe” has its customary meaning in the art, e.g., apolynucleotide which is effective to identify (e.g., by hybridization),when used in an appropriate process, the presence of a targetpolynucleotide to which it is designed. Identification can involvesimply determining presence or absence, or it can be quantitative, e.g.,in assessing amounts of a gene or gene transcript present in a sample.Probes can be useful in a variety of ways, such as for diagnosticpurposes, to identify homologs, and to detect, quantitate, or isolate apolynucleotide of the present invention in a test sample.

Assays can be utilized which permit quantification and/orpresence/absence detection of a target nucleic acid in a sample. Assayscan be performed at the single-cell level, or in a sample comprisingmany cells, where the assay is “averaging” expression over the entirecollection of cells and tissue present in the sample. Any suitable assayformat can be used, including, but not limited to, e.g., Southern blotanalysis, Northern blot analysis, polymerase chain reaction (“PCR”)(e.g., Saiki et al., Science, 241:53, 1988; U.S. Pat. Nos. 4,683,195,4,683,202, and 6,040,166; PCR Protocols: A Guide to Methods andApplications, Innis et al., eds., Academic Press, New York, 1990),reverse transcriptase polymerase chain reaction (“RT-PCR”), anchoredPCR, rapid amplification of cDNA ends (“RACF”) (e.g., Schaefer in GeneCloning and Analysis: Current Innovations, Pages 99-115, 1997), ligasechain reaction (“LCR”) (EP 320 308), one-sided PCR (Ohara et al., Proc.Natl. Acad. Sci., 86:5673-5677, 1989), indexing methods (e.g., U.S. Pat.No. 5,508,169), in situ hybridization, differential display (e.g., Lianget al., Nucl. Acid. Res., 21:3269-3275, 1993; U.S. Pat. Nos. 5,262,311,5,599,672 and 5,965,409; WO97/18454; Prashar and Weissman, Proc. Natl.Acad. Sci., 93:659-663, and U.S. Pat. Nos. 6,010,850 and 5,712,126;Welsh et al., Nucleic Acid Res., 20:49654970, 1992, and U.S. Pat. No.5,487,985) and other RNA fingerprinting techniques, nucleic acidsequence based amplification (“NASBA”) and other transcription basedamplification systems (e.g., U.S. Pat. Nos. 5,409,818 and 5,554,527; WO88/10315), polynucleotide arrays (e.g., U.S. Pat. Nos. 5,143,854,5,424,186; 5,700,637, 5,874,219, and 6,054,270; PCT WO 92/10092; PCT WO90/15070), Qbeta Replicase (PCT/US87/00880), Strand DisplacementAmplification (“SDA”), Repair Chain Reaction (“RCR”), nucleaseprotection assays, subtraction-based methods, Rapid-Scan™, etc.Additional useful methods include, but are not limited to, e.g.,template-based amplification methods, competitive PCR (e.g., U.S. Pat.No. 5,747,251), redox-based assays (e.g., U.S. Pat. No. 5,871,918),Taqman-based assays (e.g., Holland et-al., Proc. Natl. Acad, Sci.,88:7276-7280, 1991; U.S. Pat. Nos. 5,210,015 and 5,994,063), real-timefluorescence-based monitoring (e.g., U.S. Pat. No. 5,928,907), molecularenergy transfer labels (e.g., U.S. Pat. Nos. 5,348,853, 5,532,129,5,565,322, 6,030,787, and 6,117,635; Tyagi and Kramer, Nature Biotech.,14:303-309, 1996). Any method suitable for single cell analysis of geneor protein expression can be used, including in situ hybridization,immunocytochemistry, MACS, FACS, flow cytometry, etc. For single cellassays, expression products can be measured using antibodies, PCR, orother types of nucleic acid amplification (e.g., Brady et al., MethodsMol. & Cell. Biol. 2, 17-25, 1990; Eberwine et al., 1992, Proc. Natl.Acad. Sci., 89, 3010-3014, 1992; U.S. Pat. No. 5,723,290). These andother methods can be carried out conventionally, e.g., as described inthe mentioned publications.

Many of such methods may require that the polynucleotide is labeled, orcomprises a particular nucleotide type useful for detection. The presentinvention includes such modified polynucleotides that are necessary tocarry out such methods. Thus, polynucleotides can be DNA, RNA, DNA:RNAhybrids, PNA, etc., and can comprise any modification or substituentwhich is effective to achieve detection.

Detection can be desirable for a variety of different purposes,including research, diagnostic, prognostic, and forensic. For diagnosticpurposes, it may be desirable to identify the presence or quantity of apolynucleotide sequence in a sample, where the sample is obtained fromtissue, cells, body fluids, etc. In a preferred method as described inmore detail below, the present invention relates to a method ofdetecting a polynucleotide comprising, contacting a targetpolynucleotide in a test sample with a polynucleotide probe underconditions effective to achieve hybridization between the target andprobe; and detecting hybridization.

Any test sample in which it is desired to identify a polynucleotide orpolypeptide thereof can be used, including, e.g., brood, urine, saliva,stool (for extracting nucleic acid, see, e.g., U.S. Pat. No. 6,177,251),swabs comprising tissue, biopsied tissue, tissue sections, culturedcells, etc.

Detection can be accomplished in combination with polynucleotide probesfor other genes, e.g., genes which are expressed in other diseasestates, tissues, cells, such as brain, breast, heart, kidney, spleen,thymus, liver, stomach, small intestine, colon, muscle, lung, testis,placenta, pituitary, thyroid, skin, adrenal gland, pancreas, salivarygland, uterus, ovary, prostate gland, peripheral blood cells (T-cells,lymphocytes, etc.), embryo, fat, adult and embryonic stem cells,specific cell-types, such as endothelial, epithelial, myocytes, adipose,luminal epithelial, basoepithelial, myoepithelial, stromal cells, etc.

Polynucleotides can be used in wide range of methods and compositions,including for detecting, diagnosing, staging, grading, assessing,prognosticating, etc. diseases and disorders associated withdifferentially-regulated genes of the present invention, for monitoringor assessing therapeutic and/or preventative measures, in orderedarrays, etc. Any method of detecting genes and polynucleotides of SEQ IDNO 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 24, 26, 28, 30, 32, 34,36, 38, 40, 42, 44, 46, or 48 can be used; certainly, the presentinvention is not to be limited how such methods are implemented.

Along these lines, the present invention relates to methods of detectingdifferentially-regulated genes described herein in a sample comprisingnucleic acid. Such methods can comprise one or more the following stepsin any effective order, e.g., contacting said sample with apolynucleotide probe under conditions effective for said probe tohybridize specifically to nucleic acid in said sample, and detecting thepresence or absence of probe hybridized to nucleic acid in said sample,wherein said probe is a polynucleotide which is SEQ ID NO 1, 3, 5, 7, 9,11, 13, 15, 17, 19, 21, 23, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 44,46, or 48, a polynucleotide having, e.g., about 70%, 80%, 85%, 90%, 95%,99%, or more sequence identity thereto, effective or specific fragmentsthereof, or complements thereto. The detection method can be applied toany sample, e.g.; cultured primary, secondary, or established celllines, tissue biopsy, blood, urine, stool, and other bodily fluids, forany purpose.

Contacting the sample with probe can be carried out by any effectivemeans in any effective environment. It can be accomplished in a solid,liquid, frozen, gaseous, amorphous, solidified, coagulated, colloid,etc., mixtures thereof, matrix. For instance, a probe in an aqueousmedium can be contacted with a sample which is also in an aqueousmedium, or which is affixed to a solid matrix, or vice-versa.

Generally, as used throughout the specification, the term “effectiveconditions” means, e.g., the particular milieu in which the desiredeffect is achieved. Such a milieu, includes, e.g., appropriate buffers,oxidizing agents, reducing agents, pH, co-factors, temperature, ionconcentrations, suitable age and/or stage of cell (such as, inparticular part of the cell cycle, or at a particular stage whereparticular genes are being expressed) where cells are being used,culture conditions (including substrate, oxygen, carbon dioxide, etc.).When hybridization is the chosen means of achieving detection, the probeand sample can be combined such that the resulting conditions arefunctional for said probe to hybridize specifically to nucleic acid insaid sample.

The phrase “hybridize specifically” indicates that the hybridizationbetween single-stranded polynucleotides is based on nucleotide sequencecomplementarity. The effective conditions are selected such that theprobe hybridizes to a preselected and/or definite target nucleic acid inthe sample. For instance, if detection of a polynucleotide set forth inSEQ ID NO 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 24, 26, 28, 30, 32,34, 36, 38, 40, 42, 44, 46, or 48 is desired, a probe can be selectedwhich can hybridize to such target gene under high stringent conditions,without significant hybridization to other genes in the sample. Todetect homologs of a polynucleotide set forth in SEQ ID NO 1, 3, 5, 7,9, 11, 13, 15, 17, 19, 21, 23, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42,44, 46, or 48, the effective hybridization conditions can be lessstringent, and/or the probe can comprise codon degeneracy, such that ahomolog is detected in the sample.

As already mentioned, the methods can be carried out by any effectiveprocess, e.g., by Northern blot analysis, polymerase chain reaction(PCR), reverse transcriptase PCR, RACE PCR, in situ hybridization, etc.,as indicated above. When PCR based techniques are used, two or moreprobes are generally used. One probe can be specific for a definedsequence which is characteristic of a selective polynucleotide, but theother probe can be specific for the selective polynucleotide, orspecific for a more general sequence, e.g., a sequence such as polyAwhich is characteristic of mRNA, a sequence which is specific for apromoter, ribosome binding site, or other transcriptional features, aconsensus sequence (e.g., representing a functional domain). For theformer aspects, 5′ and 3′ probes (e.g., polyA, Kozak, etc.) arepreferred which are capable of specifically hybridizing to the ends oftranscripts. When PCR is utilized, the probes can also be referred to as“primers” in that they can prime a DNA polymerase reaction.

In addition to testing for the presence or absence of polynucleotides,the present invention also relates to determining the amounts at whichpolynucleotides of the present invention are expressed in sample anddetermining the differential expression of such polynucleotides insamples. Such methods can involve substantially the same steps asdescribed above for presence/absence detection, e.g., contacting withprobe, hybridizing, and detecting hybridized probe, but using morequantitative methods and/or comparisons to standards.

The amount of hybridization between the probe and target can bedetermined by any suitable methods, e.g., PCR, RT-PCR, RACE PCR,Northern blot, polynucleotide microarrays, Rapid-Scan, etc., andincludes both quantitative and qualitative measurements. For furtherdetails, see the hybridization methods described above and below.Determining by such hybridization whether the target is differentiallyexpressed (e.g., up-regulated or down-regulated) in the sample can alsobe accomplished by any effective means. For instance, the target'sexpression pattern in the sample can be compared to its pattern in aknown standard, such as in a normal tissue, or it can be compared toanother gene in the same sample. When a second sample is utilized forthe comparison, it can be a sample of normal tissue that is known not tocontain diseased cells. The comparison can be performed on samples whichcontain the same amount of RNA (such as polyadenylated RNA or totalRNA), or, on RNA extracted from the same amounts of starting tissue.Such a second sample can also be referred to as a control or standard.Hybridization can also be compared to a second target in the same tissuesample. Experiments can be performed that determine a ratio between thetarget nucleic acid and a second nucleic acid (a standard or control),e.g., in a normal tissue. When the ratio between the target and controlare substantially the same in a normal and sample, the sample isdetermined or diagnosed not to contain cells. However, if the ratio isdifferent between the normal and sample tissues, the sample isdetermined to contain cancer cells. The approaches can be combined, andone or more second samples, or second targets can be used. Any secondtarget nucleic acid can be used as a comparison, including“housekeeping” genes, such as beta-actin, alcohol dehydrogenase, or anyother gene whose expression does not vary depending upon the diseasestatus of the cell.

Methods of Identifying Polymorphisms, Mutations, etc., of aDifferentially-Regulated Gene

Polynucleotides of the present invention can also be utilized toidentify mutant alleles, SNPs, gene rearrangements and modifications,and other polymorphisms of the wild-type gene. Mutant alleles,polymorphisms, SNPs, etc., can be identified and isolated from cancersthat are known, or suspected to have, a genetic component.Identification of such genes can be carried out routinely (see, abovefor more guidance), e.g., using PCR, hybridization techniques, directsequencing, mismatch reactions (see, e.g., above), RFLP analysis, SSCP(e.g., Orita et al., Proc. Natl. Acad. Sci., 86:2766, 1992), etc., wherea polynucleotide having a sequence selected from SEQ ID NO 1, 3, 5, 7,9, 11, 13, 15, 17, 19, 21, 23, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42,44, 46, or 48 is used as a probe. The selected mutant alleles, SNPs,polymorphisms, etc., can be used diagnostically to determine whether asubject has, or is susceptible to a disorder associated with adifferentially-regulated gene, as well as to design therapies andpredict the outcome of the disorder. Methods involve, e.g., diagnosing adisorder associated with a differentially-regulated gene or determiningsusceptibility to a disorder, comprising, detecting the presence of amutation in a gene represented by a polynucleotide selected from SEQ IDNO 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 24, 26, 28, 30, 32, 34,36, 38, 40, 42, 44, 46, or 48. The detecting can be carried out by anyeffective method, e.g., obtaining cells from a subject, determining thegene sequence or structure of a target gene (using, e.g., mRNA, cDNA,genomic DNA, etc), comparing the sequence or structure of the targetgene to the structure of the normal gene, whereby a difference insequence or structure indicates a mutation in the gene in the subject.Polynucleotides can also be used to test for mutations, SNPs,polymorphisms, etc., e.g., using mismatch DNA repair technology asdescribed in U.S. Pat. No. 5,683,877; U.S. Pat. No. 5,656,430; Wu etal., Proc. Natl. Acad. Sci., 89:8779-8783, 1992.

The present invention also relates to methods of detecting polymorphismsin a differentially-regulated gene, comprising, e.g., comparing thestructure of: genomic DNA comprising all or part of said gene, mRNAcomprising all or part of said gene, cDNA comprising all or part of saidgene, or a polypeptide comprising all or part of said gene, with thestructure of said gene as set forth herein. The methods can be carriedout on a sample from any source, e.g., cells, tissues, body fluids,blood, urine, stool, hair, egg, sperm, etc.

These methods can be implemented in many different ways. For example,“comparing the structure” steps include, but are not limited to,comparing restriction maps, nucleotide sequences, amino acid sequences,RFLPs, DNAse sites, DNA methylation fingerprints (e.g., U.S. Pat. No.6,214,556), protein cleavage sites, molecular weights, electrophoreticmobilities, charges, ion mobility, etc., between a standard gene and atest gene. The term “structure” can refer to any physicalcharacteristics or configurations which can be used to distinguishbetween nucleic acids and polypeptides. The methods and instruments usedto accomplish the comparing step depends upon the physicalcharacteristics which are to be compared. Thus, various techniques arecontemplated, including, e.g., sequencing machines (both amino acid andpolynucleotide), electrophoresis, mass spectrometer (U.S. Pat. Nos.6,093,541, 6,002,127), liquid chromatography, HPLC, etc.

To carry out such methods, “all or part” of the gene or polypeptide canbe compared. For example, if nucleotide sequencing is utilized, theentire gene can be sequenced, including promoter, introns, and exons, oronly parts of it can be sequenced and compared, e.g., exon 1, exon 2,etc.

Mutagenesis

Mutated polynucleotide sequences of the present invention are useful forvarious purposes, e.g., to create mutations of the polypeptides theyencode, to identify functional regions of genomic DNA, to produce probesfor screening libraries, etc. Mutagenesis can be carried out routinelyaccording to any effective method, e.g., oligonucleotide-directed(Smith, M., Ann. Rev. Genet.19:423-463, 1985), degenerateoligonucleotide-directed (Hill et al., Method Enzymology, 155:558-568,1987), region-specific (Myers et al., Science, 229:242-246, 1985;Derbyshire et al., Gene, 46:145, 1986; Ner et al., DNA, 7:127, 1988),linker-scanning (McKnight and Kingsbury, Science, 217:316-324, 1982),directed using PCR, recursive ensemble mutagenesis (Arkin and Yourvan,Proc. Natl. Acad. Sci., 89:7811-7815, 1992), random mutagenesis (e.g.,U.S. Pat. Nos. 5,096,815; 5,198,346; and 5,223,409), site-directedmutagenesis (e.g., Walder et al., Gene, 42:133, 1986; Bauer et al.,Gene, 37:73, 1985; Craik, Bio Techniques, January 1985, 12-19; Smith etal., Genetic Engineering: Principles and Methods, Plenum Press, 1981),phage display (e.g., Lowman et al., Biochem. 30:10832-10837, 1991;Ladner et al., U.S. Pat. No. 5,223,409; Huse, WIPO Publication WO92/06204), etc. Desired sequences can also be produced by the assemblyof target sequences using mutually priming oligonucleotides (Uhlmann,Gene, 71:2940, 1988). For directed mutagenesis methods, analysis of thethree-dimensional structure of a polypeptide can be used to guide andfacilitate making mutants which effect polypeptide activity. Sites ofsubstrate-enzyme interaction or other biological activities can also bedetermined by analysis of crystal structure as determined by suchtechniques as nuclear magnetic resonance, crystallography orphotoaffinity labeling. See, for example, de Vos et al., Science255:306-312, 1992; Smith et al., J. Mol. Biol. 224:899-904, 1992;Wlodaver et al., FEBS Lett. 309:59-64, 1992.

In addition, libraries of differentially-regulated genes and fragmentsthereof can be used for screening and selection of gene variants. Forinstance, a library of coding sequences can be generated by treating adouble-stranded DNA with a nuclease under conditions where the nickingoccurs, e.g., only once per molecule, denaturing the double-strandedDNA, renaturing it to for double-stranded DNA that can includesense/antisense pairs from different nicked products, removingsingle-stranded portions from reformed duplexes by treatment with S1nuclease, and ligating the resulting DNAs into an expression vector. Bythis method, expression libraries can be made comprising “mutagenized”differentially-regulated genes. The entire coding sequence or partsthereof can be used.

Polynucleotide Expression, Polypeptides Produced Thereby, andSpecific-Binding Partners Thereto.

A polynucleotide according to the present invention can be expressed ina variety of different systems, in vitro and in vivo, according to thedesired purpose. For example, a polynucleotide can be inserted into anexpression vector, introduced into a desired host, and cultured underconditions effective to achieve expression of a polypeptide coded for bythe polynucleotide, to search for specific binding partners. Effectiveconditions include any culture conditions which are suitable forachieving production of the polypeptide by the host cell, includingeffective temperatures, pH, medium, additives to the media in which thehost cell is cultured (e.g., additives which amplify or induceexpression such as butyrate, or methotrexate if the codingpolynucleotide is adjacent to a dhfr gene), cycloheximide, celldensities, culture dishes, etc. A polynucleotide can be introduced intothe cell by any effective method including, e.g., naked DNA, calciumphosphate precipitation, electroporation, injection, DEAE-Dextranmediated transfection, fusion with liposomes, association with agentswhich enhance its uptake into cells, viral transfection. A cell intowhich a polynucleotide of the present invention has been introduced is atransformed host cell. The polynucleotide can be extrachromosomal orintegrated into a chromosome(s) of the host cell. It can be stable ortransient. An expression vector is selected for its compatibility withthe host cell. Host cells include, mammalian cells, e.g., COS, CV1, BHK,CHO, HeLa, LTK, NIH 3T3, PC-3 (CRL-1435), LNCaP (CRL-1740), CA-HPV-10(CRL-2220), PZ-HPV-7 (CRL-2221), MDA-PCa 2b (CRL-2422), 22Rv1 (CRL2505),NCI-H660 (CRL-5813), HS 804.Sk (CRL-7535), LNCaP-FGF (CRL-10995), RWPE-1(CRL-11609), RWPE-2 (CRL-11610), PWR-1E (CRL 11611), rat MAT-Ly-LuB-2(CRL-2376), and other prostate cells, insect cells, such as Sf9 (S.frugipeda) and Drosophila, bacteria, such as E. coli, Streptococcus,bacillus, yeast, such as Sacharomyces, S. cerevisiae, fungal cells,plant cells, embryonic or adult stem cells (e.g., mammalian, such asmouse or human).

Expression control sequences are similarly selected for hostcompatibility and a desired purpose, e.g., high copy number, highamounts, induction, amplification, controlled expression. Othersequences which can be employed include enhancers such as from SV40,CMV, RSV, inducible promoters, cell-type specific elements, or sequenceswhich allow selective or specific cell expression. Promoters that can beused to drive its expression, include, e.g., the endogenous promoter,MMTV, SV40, trp, lac, tac, or T7 promoters for bacterial hosts; or alphafactor, alcohol oxidase, or PGH promoters for yeast. RNA promoters canbe used to produced RNA transcripts, such as T7 or SP6. See, e.g.,Melton et al., Polynucleotide Res., 12(18):7035-7056, 1984; Dunn andStudier. J. Mol. Bio., 166:477-435, 1984; U.S. Pat. No. 5,891,636;Studier et al., Gene Expression Technology, Methods in Enzymology,85:60-89, 1987. In addition, as discussed above, translational signals(including in-frame insertions) can be included.

When a polynucleotide is expressed as a heterologous gene in atransfected cell line, the gene is introduced into a cell as describedabove, under effective conditions in which the gene is expressed. Theterm “heterologous” means that the gene has been introduced into thecell line by the “hand-of-man.” Introduction of a gene into a cell lineis discussed above. The transfected (or transformed) cell expressing thegene can be lysed or the cell line can be used intact.

For expression and other purposes, a polynucleotide can contain codonsfound in a naturally-occurring gene, transcript, or cDNA, for example,e.g., as set forth in SEQ ID NO 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21,23, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 44, 46, or 48, or it cancontain degenerate codons coding for the same amino acid sequences. Forinstance, it may be desirable to change the codons in the sequence tooptimize the sequence for expression in a desired host. See, e.g., U.S.Pat. Nos. 5,567,600 and 5,567,862.

A polypeptide according to the present invention can be recovered fromnatural sources, transformed host cells (culture medium or cells)according to the usual methods, including, detergent extraction (e.g.,non-ionic detergent, Triton X-100, CHAPS, octylglucoside, IgepalCA-630), ammonium sulfate or ethanol precipitation, acid extraction,anion or cation exchange chromatography, phosphocellulosechromatography, hydrophobic interaction chromatography, hydroxyapatitechromatography, lectin chromatography, gel electrophoresis. Proteinrefolding steps can be used, as necessary, in completing theconfiguration of the mature protein. Finally, high performance liquidchromatography (HPLC) can be employed for purification steps. Anotherapproach is express the polypeptide recombinantly with an affinity tag(Flag epitope, HA epitope, myc epitope, 6×His, maltose binding protein,chitinase, etc) and then purify by anti-tag antibody-conjugated affinitychromatography.

The present invention also relates to polypeptides corresponding to SEQID NOS 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 24, 26, 28, 30, 32,34, 36, 38, 40, 42, 44, 46, or 48, e.g., an isolated human polypeptidecomprising or having the amino acid sequence set forth in SEQ ID NOS 2,4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 25, 27, 29, 31, 33, 35, 37, 41, 43,45, or 47, an isolated human polypeptide comprising an amino acidsequence having 90%, 95%, etc., or more amino acid sequence identity tothe amino acid sequence set forth in SEQ ID NOS 2, 4, 6, 8, 10, 12, 14,16, 18, 20, 22, 25, 27, 29, 31, 33, 35, 37, 41, 43, 45, or 47. Fragmentsspecific to these polypeptides can also used, e.g., to produceantibodies or other immune responses, as competitors to its activity,etc. These fragments can be referred to as being “specific for” saidpolypeptide. The latter phrase, as already defined, indicates that thepeptides are characteristic of the polypeptide, and that the definedsequences are substantially absent from all other protein types. Suchpolypeptides can be of any size which is necessary to conferspecificity, e.g., 5, 8, 10, 12, 15, 20, or more, etc.

The present invention also relates to specific-binding partners. Theseinclude antibodies which are specific for polypeptides encoded bypolynucleotides of the present invention, as well as otherbinding-partners which interact with polynucleotides and polypeptides ofthe present invention. Protein-protein interactions between apolypeptide of the present invention and other polypeptides and bindingpartners can be identified using any suitable methods, e.g., proteinbinding assays (e.g., filtration assays, chromatography, etc.), yeasttwo-hybrid system (Fields and Song, Nature, 340: 245-247, 1989), proteinarrays, gel-shift assays, FRET (fluorescence resonance energy transfer)assays, etc. Nucleic acid interactions (e.g., protein-DNA orprotein-RNA) can be assessed using gel-shift assays, e.g., as carriedout in U.S. Pat. Nos. 6,333,407 and 5,789,538.

Antibodies, e.g., polyclonal, monoclonal, recombinant, chimeric,humanized, single-chain, Fab, and fragments thereof, can be preparedaccording to any desired method. See, also, screening recombinantimmunoglobulin libraries (e.g., Orlandi et al., Proc. Natl. Acad. Sci.,86:3833-3837, 1989; Huse et al., Science, 256:1275-1281, 1989); in vitrostimulation of lymphocyte populations; Winter and Milstein, Nature, 349:293-299, 1991. The antibodies can be IgM, IgG, subtypes, IgG2a, IgG1,etc. Antibodies, and immune responses, can also be generated byadministering naked DNA See, e.g., U.S. Pat. Nos. 5,703,055; 5,589,466;5,580,859. Antibodies can be used from any source, including, goat,rabbit, mouse, chicken (e.g., IgY; see, Duan, WO/029444 for methods ofmaking antibodies in avian hosts, and harvesting the antibodies from theeggs). An antibody specific for a polypeptide means that the antibodyrecognizes a defined sequence of amino acids within or including thepolypeptide. Other specific binding partners include, e.g., aptamers andPNA, can be prepared against specific epitopes or domains ofdifferentially regulated genes. The preparation of antibodies iswell-known to those skilled in the art. See, for example, Green et al.,Production of Polyclonal Antisera, in IMMUNOCHEMICAL PROTOCOLS (Manson,ed.), pages 1-5 (Humana Press 1992); Coligan et al., Production ofPolyclonal Antisera in Rabbits, Rats, Mice and Hamsters, in CURRENTPROTOCOLS IN IMMUNOLOGY, section 2.4.1 (1992); Kohler & Milstein, Nature256:495 (1975); Coligan et al., sections 2.5.1-2.6.7; and Harlow et al.,ANTIBODIES: A LABORATORY MANUAL, page 726 (Cold Spring Harbor Pub.1988). Antibodies can also be humanized, e.g., where they are to be usedtherapeutically.

The term “antibody” as used herein includes intact molecules as well asfragments thereof, such as Fab, F(ab′)₂, and Fv which are capable ofbinding to an epitopic determinant present in Bin1 polypeptide. Suchantibody fragments retain some ability to selectively bind with itsantigen or receptor. The term “epitope” refers to an antigenicdeterminant on an antigen to which the paratope of an antibody binds.Epitopic determinants usually consist of chemically active surfacegroupings of molecules such as amino acids or sugar side chains andusually have specific three dimensional structural characteristics, aswell as specific charge characteristics. Antibodies can be preparedagainst specific epitopes or polypeptide domains.

Antibodies which bind to a differentially-regulated polypeptide of thepresent invention can be prepared using an intact polypeptide orfragments containing small peptides of interest as the immunizingantigen. For example, it may be desirable to produce antibodies thatspecifically bind to the N- or C-terminal domains of said polypeptide.The polypeptide or peptide used to immunize an animal which is derivedfrom translated cDNA or chemically synthesized which can be conjugatedto a carrier protein, if desired. Such commonly used carriers which arechemically coupled to the immunizing peptide include keyhole limpethemocyanin (KLH), thyroglobulin, bovine serum albumin (BSA), and tetanustoxoid.

Methods of Detecting Polypeptides

Polypeptides coded for by a differentially-regulated gene of the presentinvention can be detected, visualized, determined, quantitated, etc.according to any effective method. useful methods include, e.g., but arenot limited to, immunoassays, RIA (radioimmunassay), ELISA,(enzyme-linked-immunosorbent assay), immunoflourescence, flow cytometry,histology, electron microscopy, light microscopy, in situ assays,immunoprecipitation, Western blot, etc.

Immunoassays may be carried in liquid or on biological support. Forinstance, a sample (e.g., blood, stool, urine, cells, tissue, bodyfluids, etc.) can be brought in contact with and immobilized onto asolid phase support or carrier such as nitrocellulose, or other solidsupport that is capable of immobilizing cells, cell particles or solubleproteins. The support may then be washed with suitable buffers followedby treatment with the detectably labeled differentially-regulated genespecific antibody. The solid phase support can then be washed with abuffer a second time to remove unbound antibody. The amount of boundlabel on solid support may then be detected by conventional means.

A “solid phase support or carrier” includes any support capable ofbinding an antigen, antibody, or other specific binding partner.Supports or carriers include glass, polystyrene, polypropylene,polyethylene, dextran, nylon, amylases, natural and modified celluloses,polyacrylamides, and magnetite. A support material can have anystructural or physical configuration. Thus, the support configurationmay be spherical, as in a bead, or cylindrical, as in the inside surfaceof a test tube, or the external surface of a rod. Alternatively, thesurface may be flat such as a sheet, test strip, etc. Preferred supportsinclude polystyrene beads.

One of the many ways in which gene peptide-specific antibody can bedetectably labeled is by linking it to an enzyme and using it in anenzyme immunoassay (EIA). See, e.g., Voller, A., “The Enzyme LinkedImmunosorbent Assay (ELISA),” 1978, Diagnostic Horizons 2, 1-7,Microbiological Associates Quarterly Publication, Walkersville, Md.);Voller, A. et al., 1978, J. Clin. Pathol. 31, 507-520; Butler, J. E.,1981, Meth. Enzymol. 73, 482-523; Maggio, E. (ed.), 1980, EnzymeImmunoassay, CRC Press, Boca Raton, Fla. The enzyme which is bound tothe antibody will react with an appropriate substrate, preferably achromogenic substrate, in such a manner as to produce a chemical moietythat can be detected, for example, by spectrophotometric, fluorimetricor by visual means. Enzymes that can be used to detectably label theantibody include, but are not limited to, malate dehydrogenase,staphylococcal nuclease, delta-5-steroid isomerase, yeast alcoholdehydrogenase, .alpha.-glycerophosphate, dehydrogenase, triose phosphateisomerase, horseradish peroxidase, alkaline phosphatase, asparaginase,glucose oxidase, .beta.-galactosidase, ribonuclease, urease, catalase,glucose-6-phosphate dehydrogenase, glucoamylase andacetylcholinesterase. The detection can be accomplished by colorimetricmethods that employ a chromogenic substrate for the enzyme. Detectionmay also be accomplished by visual comparison of the extent of enzymaticreaction of a substrate in comparison with similarly prepared standards.

Detection may also be accomplished using any of a variety of otherimmunoassays. For example, by radioactively labeling the antibodies orantibody fragments, it is possible to detect differentially-regulatedpeptides through the use of a radioimmunoassay (RIA). See, e.g.,Weintraub, B., Principles of Radioimmunoassays, Seventh Training Courseon Radioligand Assay Techniques, The Endocrine Society, March, 1986. Theradioactive isotope can be detected by such means as the use of a gammacounter or a scintillation counter or by autoradiography.

It is also possible to label the antibody with a fluorescent compound.When the fluorescently labeled antibody is exposed to light of theproper wave length, its presence can then be detected due tofluorescence. Among the most commonly used fluorescent labelingcompounds are fluorescein isothiocyanate, rhodamine, phycoerythrin,phycocyanin, allophycocyanin, o-phthaldehyde and fluorescamine. Theantibody can also be detectably labeled using fluorescence emittingmetals such as those in the lanthanide series. These metals can beattached to the antibody using such metal chelating groups asdiethylenetriaminepentacetic acid (DTPA) or ethyl enediaminetetraaceticacid (EDTA).

The antibody also can be detectably labeled by coupling it to achemiluminescent compound. The presence of the chemiluminescent-taggedantibody is then determined by detecting the presence of luminescencethat arises during the course of a chemical-reaction. Examples of usefulchemiluminescent labeling compounds are luminol, isoluminol, theromaticacridinium ester, imidazole, acridinium salt and oxalate ester.

Likewise, a bioluminescent compound may be used to label the antibody ofthe present invention. Bioluminescence is a type of chemiluminescencefound in biological systems in which a catalytic protein increases theefficiency of the chemiluminescent reaction. The presence of abioluminescent protein is determined by detecting the presence ofluminescence. Important bioluminescent compounds for purposes oflabeling are luciferin, luciferase and aequorin.

Tissue and Disease

The prostate is a secretory organ surrounding the neck of the bladderand urethra. Its primary function is to produce fluids and othermaterials necessary for sperm transport and maintenance. Structurally,it has both glandular and nonglandular components. The glandularcomponent is predominantly comprised of ducts and acini responsible forthe production and transport prostatic fluids. Epithelial cells are themain identifiable cell found in these regions, primarily of the basaland secretory types, but also endocrine-paracrine and transitionalepithelial. The non-glandular component contains the capsular and muscletissues, which, respectively, hold the organ together and function influid discharge. See, e.g., Histoloy for Pathologists Stemberg, S.S.,editor, Raven Press, NY, 1992, Chapter 40.

The major diseases of the prostate include, e.g., prostatic hyperplasia(BPH), prostatitis, and prostate cancer (e.g., prostaticadenocarcinoma). BPH is a benign, proliferative disease of the prostaticepithelial cells. While it may cause urinary tract obstruction in somepatients, for the most part, it is generally asymptomatic. Prostatecancer, on the other hand, is the most common form of cancer in whitemales in the United States, occurring predominantly in males over age50. The prevalence of prostate diseases, such as prostate cancer, hasmade the discovery of prostate selective markers and gene expressionpatterns of great importance.

The most common scale of assessing prostate pathology is the Gleasongrading system. See, e.g., Bostwick, Am. J. Clin. Path., 102: s38-s56,1994. Once the cancer is identified, staging can assess the size,location, and extent of the cancer. Several different staging scales arecommonly used, including stages A-D, and Tumor-Nodes-Metastases (TNM).For treatment, diagnosis, staging, etc., of prostate conditions, methodscan be carried out analogously to, and in combination with, U.S. Pat.Nos. 6,107,090; 6,057,116; 6,034,218; 6,004,267; 5,919,638; 5,882,864;5,763,202; 5,747,264; 5,688,649; 5,552,277.

In addition, the present invention relates to methods of assessing atherapeutic or preventative intervention in a subject having a prostatecancer, comprising, e.g., detecting the expression levels ofup-regulated target genes, wherein the target genes comprise a genewhich is represented by a sequence selected from SEQ ID NOS 1-47, or, agene represented by a sequence having 95% sequence identity or more to asequence selected from SEQ ID NOS 1-47. By “therapeutic or preventativeintervention,” it is meant, e.g., a drug administered a patient,surgery, radiation, chemotherapy, and other measures taken to prevent acancer or treat a cancer.

Grading, Staging, Comparing, Assessing, Methods and Compositions

The present invention also relates to methods and compositions forstaging and grading cancers. As already defined, staging relates todetermining the extent of a cancer's spread, including its size and thedegree to which other tissues, such as lymph nodes are involved in thecancer. Grading refers to the degree of a cell's retention of thecharacteristics of the tissue of its origin. A lower grade cancercomprises tumor cells that more closely resemble normal cells than amedium or higher grade cancer. Grading can be a useful diagnostic andprognostic tool. Higher grade cancers usually behave more aggressivelythan lower grade cancers. Thus, knowledge of the cancer grade, as wellas its stage, can be a significant factor in the choice of theappropriate therapeutic intervention for the particular patient, e.g.,surgery, radiation, chemotherapy, etc. Staging and grading can also beused in conjunction with a therapy to assess its efficacy, to determineprognosis, to determine effective dosages, etc.

Various methods of staging and grading cancers can be employed inaccordance with the present invention. A “cell expression profile” or“cell expression fingerprint” is a representation of the expression ofvarious different genes in a given cell or sample comprising cells.These cell expression profiles can be useful as reference standards. Thecell expression fingerprints can be used alone for grading, or incombination with other grading methods.

The present invention also relates to methods and compositions fordiagnosing a prostate cancer, or determining susceptibility to aprostate cancer, using polynucleotides, polypeptides, andspecific-binding partners of the present invention to detect, assess,determine, etc., differentially-regulated genes of the presentinvention. In such methods, the gene can serve as a marker for prostatecancer, e.g., where the gene, when mutant, is a direct cause of theprostate cancer; where the gene is affected by another gene(s) which isdirectly responsible for the prostate cancer, e.g., when the gene ispart of the same signaling pathway as the directly responsible gene;and, where the gene is chromosomally linked to the gene(s) directlyresponsible for the prostate cancer, and segregates with it. Many othersituations are possible. To detect, assess, determine, etc., a probespecific for the gene can be employed as described above and below. Anymethod of detecting and/or assessing the gene can be used, includingdetecting expression of the gene using polynucleotides, antibodies, orother specific-binding partners.

The present invention relates to methods of diagnosing a disorderassociated with prostate cancer, or determining a subject'ssusceptibility to such prostate cancer, comprising, e.g., assessing theexpression of a differentially-regulated gene in a tissue samplecomprising tissue or cells suspected of having the prostate cancer(e.g., where the sample comprises prostate). The phrase “diagnosing”indicates that it is determined whether the sample has a prostate cancercells. “Determining a subject's susceptibility to a prostate cancer”indicates that the subject is assessed for whether s/he is predisposedto get such a disease or disorder, where the predisposition is indicatedby abnormal expression of the gene (e.g., gene mutation, gene expressionpattern is not normal, etc.). Predisposition or susceptibility to adisease may result when a such disease is influenced by epigenetic,environmental, etc., factors.

By the phrase “assessing expression of a differentially-regulated gene,”it is meant that the functional status of the gene is evaluated. Thisincludes, but is not limited to, measuring expression levels of saidgene, determining the genomic structure of said gene, determining themRNA structure of transcripts from said gene, or measuring theexpression levels of polypeptide coded for by said gene. Thus, the term“assessing expression” includes evaluating the all aspects of thetranscriptional and translational machinery of the gene. For instance,if a promoter defect causes, or is suspected of causing, the disorder,then a sample can be evaluated (i.e., “assessed”) by looking (e.g.,sequencing or restriction mapping) at the promoter sequence in the gene,by detecting transcription products (e.g., RNA), by detectingtranslation product (e.g., polypeptide). Any measure of whether the geneis functional can be used, including, polypeptide, polynucleotide, andfunctional assays for the gene's biological activity.

In making the assessment, it can be useful to compare the results to anormal gene, e.g., a gene which is not associated with the disorder. Thenature of the comparison can be determined routinely, depending upon howthe assessing is accomplished. If, for example, the mRNA levels of asample is detected, then the mRNA levels of a normal can serve as acomparison, or a gene which is known not to be affected by the disorder.Methods of detecting mRNA are well known, and discussed above, e.g., butnot limited to, Northern blot analysis, polymerase chain reaction (PCR),reverse transcriptase PCR, RACE PCR, etc. Similarly, if polypeptideproduction is used to evaluate the gene, then the polypeptide in anormal tissue sample can be used as a comparison, or, polypeptide from adifferent gene whose expression is known not to be affected by thedisorder. These are only examples of how such a method could be carriedout.

The genes and polypeptides of the present invention can be used toidentify, detect, stage, determine the presence of, prognosticate,treat, study, etc., diseases and conditions of prostate as mentionedabove. The present invention relates to methods of identifying a geneticbasis for a disease or disease-susceptibility, comprising, e.g.,determining the association of prostate cancer, or prostate cancersusceptibility with a gene of the present invention. An associationbetween a disease or disease-susceptibility and nucleotide sequenceincludes, e.g., establishing (or finding) a correlation (orrelationship) between a DNA marker (e.g., gene, VNTR, polymorphism, EST,etc.) and a particular disease state. Once a relationship is identified,the DNA marker can be utilized in diagnostic tests and as a drug target.Any region of the gene can be used as a source of the DNA marker, exons,introns, intergenic regions, etc.

Human linkage maps can be constructed to establish a relationshipbetween a gene and prostate cancer. Typically, polymorphic molecularmarkers-(e.g., STRP's, SNP's, RFLP's, VNTR's) are identified within theregion, linkage and map distance between the markers is thenestablished, and then linkage is established between phenotype and thevarious individual molecular markers. Maps can be produced for anindividual family, selected populations, patient populations, etc. Ingeneral, these methods involve identifying a marker associated with thedisease (e.g., identifying a polymorphism in a family which is linked tothe disease) and then analyzing the surrounding DNA to identity the generesponsible for the phenotype. See, e.g., Kruglyak et al., Am. J. Hum.Genet., 58, 1347-1363, 1996; Matise et al., Nat. Genet., 6(4):384-90,1994.

Assessing the effects of therapeutic and preventative interventions(e.g., administration of a drug, chemotherapy, radiation, etc.) onprostate cancer is a major effort in drug discovery, clinical medicine,and pharmacogenomics. The evaluation of therapeutic and preventativemeasures, whether experimental or already in clinical use, has broadapplicability, e.g., in clinical trials, for monitoring the status of apatient, for analyzing and assessing animal models, and in any scenarioinvolving cancer treatment and prevention. Analyzing the expressionprofiles of polynucleotides of the present invention can be utilized asa parameter by which interventions are judged and measured. Treatment ofa disorder can change the expression profile in some manner which isprognostic or indicative of the drug's effect on it. Changes in theprofile can indicate, e.g., drug toxicity, return to a normal level,etc. Accordingly, the present invention also relates to methods ofmonitoring or assessing a therapeutic or preventative measure (e.g.,chemotherapy, radiation, anti-neoplastic drugs, antibodies, etc.) in asubject having prostate cancer, or, susceptible to such a disorder,comprising, e.g., detecting the expression levels of one or moredifferentially-regulated genes of the present invention. A subject canbe a cell-based assay system; non-human animal model, human patient,etc. Detecting can be accomplished as described for the methods aboveand below. By “therapeutic or preventative intervention,” it is meant,e.g., a drug administered to a patient, surgery, radiation,chemotherapy, and other measures taken to prevent, treat, or diagnoseprostate cancer.

Expression can be assessed in any sample comprising any tissue or celltype, body fluid, etc., as discussed for other methods of the presentinvention, including cells from prostate can be used, or cells derivedfrom prostate. By the phrase “cells derived from prostate,” it is meantthat the derived cells originate from prostate, e.g., when metastasisfrom a primary tumor site has occurred, when a progenitor-type orpluripotent cell gives rise to other cells, etc.

The present invention-also relates to methods of using binding partners,such as antibodies, to deliver active agents to the prostate for avariety of different purposes, including, e.g., for diagnostic,therapeutic (e.g., to treat prostate cancer), and research purposes.Methods can involve delivering or administering an active agent toprostate, comprising, e.g., administering to a subject in need thereof,an effective amount of an active agent coupled to a binding partnerspecific for human polypeptide, wherein said binding partner iseffective to deliver said active agent specifically to prostate.

Any type of active agent can be used in combination with a bindingpartner, including, therapeutic, cytotoxic, cytostatic,chemotherapeutic, anti-neoplastic, anti-proliferative, anti-biotic,etc., agents. A chemotherapeutic agent can be, e.g., DNA-interactiveagent, alkylating agent, antimetabolite, tubulin-interactive agent,hormonal agent, hydroxyurea, Cisplatin, Cyclophosphamide, Altretamine,Bleomycin, Dactinomycin, Doxorubicin, Etoposide, Teniposide, paclitaxel,cytoxan, 2-methoxycarbonylaminobenzimidazole, Plicamycin, Methotrexate,Fluorouracil, Fluorodeoxyuridin, CB3717, Azacitidine, Floxuridine,Mercapyopurine, 6-Thioguanine, Pentostatin, Cytarabine, Fludarabine,etc. Agents can also be contrast agents useful in imaging technology,e.g., X-ray, CT, CAT, MRI, ultrasound, PET, SPECT, and scintographic.

An active agent can be associated in any manner with a [GENE] bindingpartner which is effective to achieve its delivery specifically to thetarget. Specific delivery or targeting indicates that the agent isprovided to the prostate, without being substantially provided to othertissues. The association of the active agent and the binding partner(“coupling) can be direct, e.g., through chemical bonds between thebinding partner and the agent, or, via a linking agent, or theassociation can be less direct, e.g., where the active agent is in aliposome, or other carrier, and the binding partner is associated withthe liposome surface. In such case, the binding partner can be orientedin such a way that it is able to bind to a polypeptide on the cellsurface. Methods for delivery of DNA via a cell-surface receptor isdescribed, e.g., in U.S. Pat. No. 6,339,139.

Identifying Agent Methods

The present invention also relates to methods of identifying agents, andthe agents themselves, which modulate prostate cancer genes. Theseagents can be used to modulate the biological activity of thepolypeptide encoded for the gene, or the gene, itself. Agents whichregulate the gene or its product are useful in variety of differentenvironments, including as medicinal agents to treat or preventdisorders associated with prostate cancer genes and as research reagentsto modify the function of tissues and cell.

Methods of identifying agents generally comprise steps in which an agentis placed in contact with the gene, transcription product, translationproduct, or other target, and then a determination is performed toassess whether the agent “modulates” the target. The specific methodutilized will depend upon a number of factors, including, e.g., thetarget (i.e., is it the gene or polypeptide encoded by it), theenvironment (e.g., in vitro or in vivo), the composition of the agent,etc.

For modulating the expression of a prostate cancer gene, a method cancomprise, in any effective order, one or more of the following steps,e.g., contacting a prostate cancer gene (e.g., in a cell population)with a test agent under conditions effective for said test agent tomodulate the expression of the prostate cancer, and determining whethersaid test agent modulates said gene. An agent can modulate expression ofa gene at any level, including transcription, translation, and/orperdurance of the nucleic acid (e.g., degradation, stability, etc.) inthe cell.

For modulating the biological activity of prostate cancer polypeptides,a method can comprise, in any effective order, one or more of thefollowing steps, e.g., contacting a polypeptide (e.g., in a cell,lysate, or isolated) with a test agent under conditions effective forsaid test agent to modulate the biological activity of said polypeptide,and determining whether said test agent modulates said biologicalactivity.

Contacting a gene or polypeptide with the test agent can be accomplishedby any suitable method and/or means that places the agent in a positionto functionally control its expression or biological activity.Functional control indicates that the agent can exert its physiologicaleffect on the gene or polypeptide through whatever mechanism it works.The choice of the method and/or means can depend upon the nature of theagent and the condition and type of environment in which the gene orpolypeptide is presented, e.g., lysate, isolated, or in a cellpopulation (such as, in vivo, in vitro, organ explants, etc.). Forinstance, if the cell population is an in vitro cell culture, the agentcan be contacted with the cells by adding it directly into the culturemedium. If the agent cannot dissolve readily in an aqueous medium, itcan be incorporated into liposomes, or another lipophilic carrier, andthen administered to the cell culture. Contact can also be facilitatedby incorporation of agent with carriers and delivery molecules andcomplexes, by injection, by infusion, etc.

Agents can be directed to, or targeted to, any part of the polypeptidewhich is effective for modulating it. For example, agents, such asantibodies and small molecules, can be targeted to cell-surface,exposed, extracellular, ligand binding, functional, etc., domains of thepolypeptide. Agents can also be directed to intracellular regions anddomains, e.g., regions where the polypeptide couples or interacts withintracellular or intramembrane binding partners.

After the agent has been administered in such a way that it can gainaccess to the gene or polypeptide, it can be determined whether the testagent modulates the gene or polypeptide expression or biologicalactivity. Modulation can be of any type, quality, or quantity, e.g.,increase, facilitate, enhance, up-regulate, stimulate, activate,amplify, augment, induce, decrease, down-regulate, diminish, lessen,reduce, etc. The modulatory quantity can also encompass any value, e.g.,1%, 5%, 10%, 50%, 75%, 1-fold, 2-fold, 5-fold, 10-fold, 100-fold, etc.To modulate gene expression means, e.g., that the test agent has aneffect on its expression, e.g., to effect the amount of transcription,to effect RNA splicing, to effect translation of the RNA intopolypeptide, to effect RNA or polypeptide stability, to effectpolyadenylation or other processing of the RNA, to effectpost-transcriptional or post-translational processing, etc. To modulatebiological activity means, e.g., that a functional activity of thepolypeptide is changed in comparison to its normal activity in theabsence of the agent. This effect includes, increase, decrease, block,inhibit, enhance, etc.

A test agent can be of any molecular composition e.g., chemicalcompounds, biomolecules, such as polypeptides, lipids, nucleic acids(e.g., antisense to a polynucleotide sequence selected from SEQ ID NO 1,3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 24, 26, 28, 30, 32, 34, 36, 38,40, 42, 44, 46, or 48), carbohydrates, antibodies, ribozymes,double-stranded RNA, aptamers, etc. For example, if a polypeptide to bemodulated is a cell-surface molecule, a test agent can be an antibodythat specifically recognizes it and, e.g., causes the polypeptide to beinternalized, leading to its down regulation on the surface of the cell.Such an effect does not have to be permanent, but can require thepresence of the antibody to continue the down-regulatory effect.Antibodies can also be used to modulate the biological activity apolypeptide in a lysate or other cell-free form. Antisense can also beused as test agents to modulate gene expression.

Markers

The polynucleotides of the present invention can be used with othermarkers, especially prostate and prostate cancer markers to identity,detect, stage, diagnosis, determine, prognosticate, treat, etc., tissue,diseases and conditions, etc, of the prostate. Markers can bepolynucleotides, polypeptides, antibodies, ligands, specific bindingpartners, etc.

A number of genes and gene products have been identified which areassociated with prostate cancer metastasis and/or progression, e.g.,PSA, KAII (shows decreased expression in metastatic cells; Dong et al.,Science, 268:884-6, 1995), D44 isoforms (differentially-regulated duringcarcinoma progression; Noordzij et al., Clin. Cancer Res., 3:805-15,1997), p53 (Effert et al., J. Urol., 150:257-61, 1993), Rb, CDKN2,E-cadherin, PTEN (Hamilton et al., Br. J. Cancer, 82:1671-6, 2000; Donget al., Clin. Cancer Res., 7:304-308, 2001), bcl-2, prostatic acidphosphatase (PAP), prostate specific membrane antigen (e.g., U.S. Pat.Nos. 5,538,866 and 6,107,090), Smad3 (e.g., Kang et al., Proc. Natl.Acad. Sci., 98:3018-3023, 2061), TGF-beta, and other oncogenes and tumorsuppressor genes. See, also, Myers and Grizzle, Eur. Urol., 30:153-166,1996, for other biomarkers associated with prostatic carcinoma, such asPCNA, p185-erbB-2, p180erbB-3, TAG-72, nm23-H1 and FASE. Such markerscan be used in combination with the methods of the present invention tofacilitate identifying, grading, staging, prognostication, etc, ofconditions and diseases of the prostate.

Therapeutics

Selective polynucleotides, polypeptides, and specific-binding partnersthereto, can be utilized in therapeutic applications, especially totreat prostate cancer. Useful methods include, but are not limited to,immunotherapy (e.g., using specific-binding partners to polypeptides),vaccination (e.g., using a selective polypeptide or a naked DNA encodingsuch polypeptide), protein or polypeptide replacement therapy, genetherapy (e.g., germ-line correction, antisense), etc.

Various immunotherapeutic approaches can be used. For instance,unlabeled antibody that specifically recognizes a tissue-specificantigen can be used to stimulate the body to destroy or attack thecancer, to cause down-regulation, to produce complement-mediated lysis,to inhibit cell growth, etc., of target cells which display the antigen,e.g., analogously to how c-erbB-2 antibodies are used to treat breastcancer. In addition, antibody can be labeled or conjugated to enhanceits deleterious effect, e.g., with radionuclides and other energyemitting entitities, toxins, such as ricin, exotoxin A (ETA), anddiphtheria, cytotoxic or cytostatic agents, immunomodulators,chemotherapeutic agents, etc. See, e.g., U.S. Pat. No. 6,107,090.

An antibody or other specific-binding partner can be conjugated to asecond molecule, such as a cytotoxic agent, and used for targeting thesecond molecule to a tissue-antigen positive cell (Vitetta, E. S. etal., 1993, Immunotoxin therapy, in DeVita, Jr., V. T. et al., eds,Cancer: Principles and Practice of Oncology, 4th ed., J. B. LippincottCo., Philadelphia, 2624-2636). Examples of cytotoxic agents include, butare not limited to, antimetabolites, alkylating agents, anthracyclines,antibiotics, anti-mitotic agents, radioisotopes and chemotherapeuticagents. Further examples of cytotoxic agents include, but are notlimited to ricin, doxorubicin, daunorubicin, taxol, ethidium bromide,mitomycin, etoposide, tenoposide, vincristine, vinblastine, coichicine,dihydroxy anthracin dione, actinomycin D, 1-dehydrotestosterone,diptheria toxin, Pseudomonas exotoxin (PE) A, PE40, abrin, elongationfactor-2 and glucocorticoid. Techniques for conjugating therapeuticagents to antibodies are well.

In addition to immunotherapy, polynucleotides and polypeptides can beused as targets for non-immunotherapeutic applications, e.g., usingcompounds which interfere with function, expression (e.g., antisense asa therapeutic agent), assembly, etc. RNA interference can be used invivtro and in vivo to silence differentially-expressed genes when itsexpression contributes to a disease (but also for other purposes, e.g.,to identify the gene's function to change a developmental pathway of acell, etc.). See, e.g., Sharp and Zamore, Science, 287:2431-2433, 2001;Grishok et al., Science, 287:2494, 2001.

Delivery of therapeutic agents can be achieved according to anyeffective method, including, liposomes, viruses, plasmid vectors,bacterial delivery systems, orally, systemically, etc. Therapeuticagents of the present invention can be administered in any form by anyeffective route, including, e.g., oral, parenteral, enteral,intraperitoneal, topical, transdermal (e.g., using any standard patch),ophthalmic, nasally, local, non-oral, such as aerosal, inhalation,subcutaneous, intramuscular, buccal, sublingual, rectal, vaginal,intravenous, intra-arterial, and intrathecal, etc. They can beadministered alone, or in combination with any ingredient(s), active orinactive.

In addition to therapeutics, per se, the present invention also relatesto methods of treating prostate cancer showing altered expression ofdifferentially-regulated genes, such as SEQ ID NO 1, 3, 5, 7, 9, 11, 13,15, 17, 19, 21, 23, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 44, 46, or48, comprising, e.g., administering to a subject in need thereof atherapeutic agent which is effective for regulating expression of saidgenes and/or which is effective in treating said disease. The term“treating” is used conventionally, e.g., the management or care of asubject for the purpose of combating, alleviating, reducing, relieving,improving the condition of, etc., of a disease or disorder. By thephrase “altered expression,” it is meant that the disease is associatedwith a mutation in the gene, or any modification to the gene (orcorresponding product) which affects its normal function. Thus,expression of a differentially-regulated gene refers to, e.g.,transcription, translation, splicing, stability of the mRNA or proteinproduct, activity of the gene product, differential expression, etc.

Any agent which “treats” the disease can be used. Such an agent can beone which regulates the expression of the gene. Expression refers to thesame acts already mentioned, e.g. transcription, translation, splicing,stability of the mRNA or protein product, activity of the gene product,differential expression, etc. For instance, if the condition was aresult of a complete deficiency of the gene product, administration ofgene product to a patient would be said to treat the disease andregulate the gene's expression. Many other possible situations arepossible, e.g., where the gene is aberrantly expressed, and thetherapeutic agent regulates the aberrant expression by restoring itsnormal expression pattern.

Antisense

Antisense polynucleotide (e.g., RNA) can also be prepared from apolynucleotide according to the present invention, preferably ananti-sense to a sequence of SEQ ID NO 1, 3, 5, 7, 9, 11, 13, 15, 17, 19,21, 23, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 44, 46, or 48. Antisensepolynucleotide can be used in various ways, such as to regulate ormodulate expression of the polypeptides they encode, e.g., inhibit theirexpression, for in situ hybridization, for therapeutic purposes, formaking targeted mutations (in vivo, triplex, etc.) etc. For guidance onadministering and designing anti-sense, see, e.g., U.S. Pat. Nos.6,200,960, 6,200,807, 6,197,584, 6,190,869, 6,190,661, 6,187,587,6,168,950, 6,153,595, 6,150,162, 6,133,246, 6,117,847, 6,096,722,6,087,343, 6,040,296, 6,005,095, 5,998,383, 5,994,230, 5,891,725,5,885,970, and 5,840,708. An antisense polynucleotides can be operablylinked to an expression control sequence. A total length of about 35 bpcan be used in cell culture with cationic liposomes to facilitatecellular uptake, but for in vivo use, preferably shorteroligonucleotides are administered, e.g. 25 nucleotides.

Antisense polynucleotides can comprise modified, nonnaturally-occurringnucleotides and linkages between the nucleotides (e.g., modification ofthe phosphate-sugar backbone; methyl phosphonate, phosphorothioate, orphosphorodithioate linkages; and 2′-O-methyl ribose sugar units), e.g.,to enhance in vivo or in vitro stability, to confer nuclease resistance,to modulate uptake, to modulate cellular distribution andcompartmentalization, etc. Any effective nucleotide or modification canbe used, including those already mentioned, as known in the art, etc.,e.g., disclosed in U.S. Pat. Nos. 6,133,438; 6,127,533; 6,124,445;6,121,437; 5,218,103 (e.g., nucleoside thiophosphoramidites); 4,973,679;Sproat et al., “2′-O-Methyloligoribonucleotides: synthesis andapplications,” Oligonucleotides and Analogs A Practical Approach,Eckstein (ed.), IRL Press, Oxford, 1991, 49-86; Iribarren et al.,“2′O-Alkyl Oligoribonucleotides as Antisense Probes,” Proc. Natl. Acad.Sci. USA, 1990, 87, 7747-7751; Cotton et al., “2′-O-methyl, 2′-O-ethyloligoribonucleotides and phosphorothioate oligodeoxyribonucleotides asinhibitors of the in vitro U7 snRNP-dependent mRNA processing event,”Nucl. Acids Res., 1991, 19, 2629-2635.

Arrays

The present invention also relates to an ordered array of polynucleotideprobes and specific-binding partners (e.g., antibodies) for detectingthe expression of differentially-regulated genes in a sample,comprising, one or more polynucleotide probes or specific bindingpartners associated with a solid support, wherein each probe is specificfor said genes, and the probes comprise a nucleotide sequence of SEQ IDNO 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 24, 26, 28, 30, 32, 34,36, 38, 40, 42, 44, 46, or 48 which is specific for said gene, anucleotide sequence having sequence identity to SEQ ID NO 1, 3, 5, 7, 9,11, 13, 15, 17, 19, 21, 23, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 44,46, or 48 which is specific for said gene or polynucleotide, orcomplements thereto, or a specific-binding partner which is specific forsaid genes.

The phrase “ordered array” indicates that the probes are arranged in anidentifiable or position-addressable pattern, e.g., such as the arraysdisclosed in U.S. Pat. Nos. 6,156,501, 6,077,673, 6,054,270, 5,723,320,5,700,637, WO09919711, WO00023803. The probes are associated with thesolid support in any effective way. For instance, the probes can bebound to the solid support, either by polymerizing the probes on thesubstrate, or by attaching a probe to the substrate. Association can be,covalent, electrostatic, noncovalent, hydrophobic, hydrophilic,noncovalent, coordination, adsorbed, absorbed, polar, etc. When fibersor hollow filaments are utilized for the array, the probes can fill thehollow orifice, be absorbed into the solid filament, be attached to thesurface of the orifice, etc. Probes can be of any effective size,sequence identity, composition, etc., as already discussed.

Ordered arrays can further comprise polynucleotide probes orspecific-binding partners which are specific for other genes, includinggenes specific for prostate or disorders associated with prostate, suchas prostate cancer.

Transgenic Animals

The present invention also relates to transgenic animals comprisingdifferentially-regulated genes, and homologs thereof, of the presentinvention. Such genes, as discussed in more detail below, include, butare not limited to, functionally-disrupted genes, mutated genes,ectopically or selectively-expressed genes, inducible or regulatablegenes, etc. These transgenic animals can be produced according to anysuitable technique or method, including homologous recombination,mutagenesis (e.g., ENU, Rathkolb et al., Exp. Physiol., 85(6):635-644,2000), and the tetracycline-regulated gene expression system (e.g., U.S.Pat. No. 6,242,667). The term “gene” as used herein includes any part ofa gene, i.e., regulatory sequences, promoters, enhancers, exons,introns, coding sequences, etc. The nucleic acid present in theconstruct or transgene can be naturally-occurring wild-type,polymorphic, or mutated. Where the animal is a non-human animal, itshomolog can be used instead. Such animals can be susceptible to prostatecancer.

Along these lines, polynucleotides of the present invention can be usedto create transgenic animals, e.g. a non-human animal, comprising atleast one cell whose genome comprises a functional disruption of adifferentially-regulated gene, or a homolog thereof (e.g., when a mouseis used, the mouse homolog corresponding to the gene can be engineered).By the phrases “functional disruption” or “functionally disrupted,” itis meant that the gene does not express a biologically-active product.It can be substantially deficient in at least one functional activitycoded for by the gene. Expression of a polypeptide can be substantiallyabsent, i.e., essentially undetectable amounts are made. However,polypeptide can also be made, but which is deficient in activity, e.g.,where only an amino-terminal portion of the gene product is produced.

The transgenic animal can comprise one or more cells. When substantiallyall its cells contain the engineered gene, it can be referred to as atransgenic animal “whose genome comprises” the engineered gene. Thisindicates that the endogenous gene loci of the animal has been modifiedand substantially all cells contain such modification.

Functional disruption of the gene can be accomplished in any effectiveway, including, e.g., introduction of a stop codon into any part of thecoding sequence such that the resulting polypeptide is biologicallyinactive (e.g., because it lacks a catalytic domain, a ligand bindingdomain, etc.), introduction of a mutation into a promoter or otherregulatory sequence that is effective to turn it off, or reducetranscription of the gene, insertion of an exogenous sequence into thegene which inactivates it (e.g., which disrupts the production of abiologically-active polypeptide or which disrupts the promoter or othertranscriptional machinery), deletion of sequences from the adifferentially-regulated gene, etc. Examples of transgenic animalshaving functionally disrupted genes are well known, e.g., as describedin U.S. Pat. Nos. 6,239,326, 6,225,525, 6,207,878, 6,194,633, 6,187,992,6,180,849, 6,177,610, 6,100,445, 6,087,555, 6,080,910, 6,069,297,6,060,642, 6,028,244, 6,013,858,5,981,830, 5,866,760, 5,859,314,5,850,004, 5,817,912, 5,789,654, 5,777,195, and 5,569,824. A transgenicanimal which comprises the functional disruption can also be referred toas a “knock-out” animal, since the biological activity of its adifferentially-regulated gene has been “knocked-out.” Knock-outs can behomozygous or heterozygous.

For creating functional disrupted genes, and other gene mutations,homologous recombination technology is of special interest since itallows specific regions of the genome to be targeted. Using homologousrecombination methods, genes can be specifically-inactivated, specificmutations can be introduced, and exogenous sequences can be introducedat specific sites. These methods are well known in the art, e.g., asdescribed in the patents above. See, also, Robertson, Biol. Reproduc.,44(2):238-245, 1991. Generally, the genetic engineering is performed inan embryonic stem (ES) cell, or other pluripotent cell line (e.g., adultstem cells, EG cells), and that genetically-modified cell (or nucleus)is used to create a whole organism. Nuclear transfer can be used incombination with homologous recombination technologies.

For example, a differentially-regulated gene locus can be disrupted inmouse ES cells using a positive-negative selection method (e.g., Mansouret al., Nature, 336:348-352, 1988). In this method, a targeting vectorcan be constructed which comprises a part of the gene to be targeted. Aselectable marker, such as neomycin resistance genes, can be insertedinto a a differentially-regulated gene exon present in the targetingvector, disrupting it. When the vector recombines with the ES cellgenome, it disrupts the function of the gene. The presence in the cellof the vector can be determined by expression of neomycin resistance.See, e.g., U.S. Pat. No. 6,239,326. Cells having at least onefunctionally disrupted gene can be used to make chimeric and germlineanimals, e.g., animals having somatic and/or germ cells comprising theengineered gene. Homozygous knock-out animals can be obtained frombreeding heterozygous knock-out animals. See, e.g., U.S. Pat. No.6,225,525.

A transgenic animal, or animal cell, lacking one or more functionaldifferentially-regulated genes can be useful in a variety ofapplications, including, as an animal model for cancer, for drugscreening assays, as a source of tissues deficient in said geneactivity, and any of the utilities mentioned in any issued U.S. Patenton transgenic animals, including, U.S. Pat. Nos. 6,239,326, 6,225,525,6,207,878, 6,194,633, 6,187,992, 6,180,849, 6,177,610, 6,100,445,6,087,555,6,080,910, 6,069,297, 6,060,642, 6,028,244, 6,013,858,5,981,830, 5,866,760, 5,859,314, 5,850,004, 5,817,912, 5,789,654,5,777,195, and 5,569,824.

The present invention also relates to non-human, transgenic animal whosegenome comprises recombinant a differentially-regulated gene nucleicacid operatively linked to an expression control sequence effective toexpress said coding sequence, e.g., in prostate. such a transgenicanimal can also be referred to as a “knock-in” animal since an exogenousgene has been introduced, stably, into its genome.

A recombinant a differentially-regulated gene nucleic acid refers to agene which has been introduced into a target host cell and optionallymodified, such as cells derived from animals, plants, bacteria, yeast,etc. A recombinant a differentially-regulated gene includes completelysynthetic nucleic acid sequences, semi-synthetic nucleic acid sequences,sequences derived from natural sources, and chimeras thereof. “Operablelinkage” has the meaning used through the specification, i.e., placed ina functional relationship with another nucleic acid. When a gene isoperably linked to an expression control sequence, as explained above,it indicates that the gene (e.g., coding sequence) is joined to theexpression control sequence (e.g., promoter) in such a way thatfacilitates transcription and translation of the coding sequence. Asdescribed above, the phrase “genome” indicates that the genome of thecell has been modified. In this case, the recombinant adifferentially-regulated gene has been stably integrated into the genomeof the animal. The a differentially-regulated gene nucleic acid inoperable linkage with the expression control sequence can also bereferred to as a construct or transgene.

Any expression control sequence can be used depending on the purpose.For instance, if selective expression is desired, then expressioncontrol sequences which limit its expression can be selected. Theseinclude, e.g., tissue or cell-specific promoters, introns, enhancers,etc. For various methods of cell and tissue-specific expression, see,e.g., U.S. Pat. Nos. 6,215,040, 6,210,736, and 6,153,427. These alsoinclude the endogenous promoter, i.e., the coding sequence can beoperably linked to its own promoter. Inducible and regulatable promoterscan also be utilized.

The present invention also relates to a transgenic animal which containsa functionally disrupted and a transgene stably integrated into theanimals genome. Such an animal can be constructed using combinations anyof the above- and below-mentioned methods. Such animals have any of theaforementioned uses, including permitting the knock-out of the normalgene and its replacement with a mutated gene. Such a transgene can beintegrated at the endogenous gene locus so that the functionaldisruption and “knock-in” are carried out in the same step.

In addition to the methods mentioned above, transgenic animals can beprepared according to known methods, including, e.g., by pronuclearinjection of recombinant genes into pronuclei of 1-cell embryos,incorporating an artificial yeast chromosome into embryonic stem cells,gene targeting methods, embryonic stem cell methodology, cloningmethods, nuclear transfer methods. See, also, e.g., U.S. Pat. Nos.4,736,866; 4,873,191; 4,873,316; 5,082,779; 5,304,489; 5,174,986;5,175,384; 5,175,385; 5,221,778; Gordon et al., Proc. Natl. Acad. Sci.,77:7380-7384, 1980; Palmiter et al., Cell, 41:343-345, 1985; Palmiter etal., Ann. Rev. Genet., 20:465-499, 1986; Askew et al., Mol. Cell. Bio.,13:4115-4124, 1993; Games et al. Nature, 373:523-527, 1995; Valanciusand Smithies, Mol. Cell. Bio., 11: 1402-1408, 1991; Stacey et al., Mol.Cell. Bio., 14:1009-1016, 1994; Hasty et al., Nature, 350:243-246, 1995;Rubinstein et al., Nucl. Acid Res., 21:2613-2617,1993; Cibelli et al.,Science, 280:1256-1258, 1998. For guidance on recombinase excisionsystems, see, e.g., U.S. Pat. Nos. 5,626,159, 5,527,695, and 5,434,066.See also, Orban, P. C., et al., “Tissue- and Site-Specific DNARecombination in Transgenic Mice,” Proc. Natl. Acad. Sci. USA,89:6861-6865 (1992); O'Gorman, S., et al., “Recombinase-Mediated GeneActivation and Site-Specific Integration in Mammalian Cells,” Science,251:1351-1355 (1991); Sauer, B., et al., “Cre-stimulated recombinationat loxP-Containing DNA sequences placed into the mammalian genome,”Polynucleotides Research, 17(1):147-161 (1989); Gagneten, S. et al.(1997) Nucl. Acids Res. 25:3326-3331; Xiao and Weaver (1997) Nucl. AcidsRes. 25:2985-2991; Agah, R. et al. (1997) J. Clin. Invest. 100:169-179;Barlow, C. et al. (1997) Nucl. Acids Res. 25:2543-2545; Araki, K. et al.(1997) Nucl. Acids Res. 25:868-872; Mortensen, R. N. et al. (1992) Mol.Cell. Biol. 12:2391-2395 (G418 escalation method); Lakhlani, P. P. etal. (1997) Proc. Natl. Acad. Sci. USA 94:9950-9955 (“hit and run”);Westphal and Leder (1997) Curr. Biol. 7:530-533 (transposon-generated“knock-out” and “knock-in”); Templeton, N. S. et al. (1997) Gene Ther.4:700-709 (methods for efficient gene targeting, allowing for a highfrequency of homologous recombination events, e.g., without selectablemarkers); PCT International Publication WO 93/22443(functionally-disrupted).

A polynucleotide according to the present invention can be introducedinto any non-human animal, including a non-human mammal, mouse (Hogan etal., Manipulating the Mouse Embryo: A Laboratory Manual. Cold SpringHarbor Laboratory, Cold Spring Harbor, N.Y., 1986), pig (Hammer et al.,Nature, 315:343-345, 1985), sheep (Hammer et al., Nature, 315:343-345,1985), cattle, rat, or primate. See also, e.g., Church, 1987, Trends inBiotech. 5:13-19; Clark et al., Trends in Biotech. 5:20-24, 1987); andDePamphilis et al., BioTechniques, 6:662-680, 1988. Transgenic animalscan be produced by the methods described in U.S. Pat. No. 5,994,618, andutilized for any of the utilities described therein.

Database

The present invention also relates to electronic forms ofpolynucleotides, polypeptides, etc., of the present invention, includingcomputer-readable medium (e.g., magnetic, optical, etc., stored in anysuitable format, such as flat files or hierarchical files) whichcomprise such sequences, or fragments thereof, e-commerce-related means,etc. Along these lines, the present invention relates to methods ofretrieving gene sequences from a computer-readable medium, comprising,one or more of the following steps in any effective order, e.g.,selecting a cell or gene expression profile, e.g., a profile thatspecifies that said gene is differentially expressed in prostate cancer,and retrieving said differentially expressed gene sequences, where thegene sequences consist of the genes represented by SEQ ID NO 1, 3, 5, 7,9, 11, 13, 15, 17, 19, 21, 23, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42,44, 46, or 48.

A “gene expression profile” means the list of tissues, cells, etc., inwhich a defined gene is expressed (i.e, transcribed and/or translated).A “cell expression profile” means the genes which are expressed in theparticular cell type. The profile can be a list of the tissues in whichthe gene is expressed, but can include additional information as well,including level of expression (e.g., a quantity as compared ornormalized to a control gene), and information on temporal (e.g., atwhat point in the cell-cycle or developmental program) and spatialexpression. By the phrase “selecting a gene or cell expression profile,”it is meant that a user decides what type of gene or cell expressionpattern he is interested in retrieving, e.g., he may require that thegene is differentially expressed in a tissue, or he may require that thegene is not expressed in blood, but must be expressed in prostatecancer. Any pattern of expression preferences may be selected. Theselecting can be performed by any effective method. In general,“selecting” refers to the process in which a user forms a query that isused to search a database of gene expression profiles. The step ofretrieving involves searching for results in a database that correspondto the query set forth in the selecting step. Any suitable algorithm canbe utilized to perform the search query, including algorithms that lookfor matches, or that perform optimization between query and data. Thedatabase is information that has been stored in an appropriate storagemedium, having a suitable computer-readable format. Once results areretrieved, they can be displayed in any suitable format, such as HTML.

For instance, the user may be interested in identifying genes that aredifferentially expressed in a prostate cancer. He may not care whethersmall amounts of expression occur in other tissues, as long as suchgenes are not expressed in peripheral blood lymphocytes. A query isformed by the user to retrieve the set of genes from the database havingthe desired gene or cell expression profile. Once the query is inputtedinto the system, a search algorithm is used to interrogate the database,and retrieve results.

Advertising, Licensing, etc., Methods

The present invention also relates to methods of advertising, licensing,selling, purchasing, brokering, etc., genes, polynucleotides,specific-binding partners, antibodies, etc., of the present invention.Methods can comprises, e.g., displaying a differentially-regulated gene,a differentially-regulated gene polypeptide, or antibody specific for adifferentially-regulated gene in a printed or computer-readable medium(e.g., on the Web or Internet), accepting an offer to purchase saidgene, polypeptide, or antibody.

Other

A polynucleotide, probe, polypeptide, antibody, specific-bindingpartner, etc., according to the present invention can be isolated. Theterm “isolated” means that the material is in a form in which it is notfound in its original environment or in nature, e.g., more concentrated,more purified, separated from component, etc. An isolated polynucleotideincludes, e.g., a polynucleotide having the sequenced separated from thechromosomal DNA found in a living animal, e.g., as the complete gene, atranscript, or a cDNA. This polynucleotide can be part of a vector orinserted into a chromosome (by specific gene-targeting or by randomintegration at a position other than its normal position) and still beisolated in that it is not in a form that is found in its naturalenvironment. A polynucleotide, polypeptide, etc., of the presentinvention can also be substantially purified. By substantially purified,it is meant that polynucleotide or polypeptide is separated and isessentially free from other polynucleotides or polypeptides, i.e., thepolynucleotide or polypeptide is the primary and active constituent. Apolynucleotide can also be a recombinant molecule. By “recombinant,” itis meant that the polynucleotide is an arrangement or form which doesnot occur in nature. For instance, a recombinant molecule comprising apromoter sequence would not encompass the naturally-occurring gene, butwould include the promoter operably linked to a coding sequence notassociated with it in nature, e.g., a reporter gene, or a truncation ofthe normal coding sequence.

The term “marker” is used herein to indicate a means for detecting orlabeling a target. A marker can be a polynucleotide (usually referred toas a “probe”), polypeptide (e.g., an antibody conjugated to a detectablelabel), PNA, or any effective material.

The topic headings set forth above are meant as guidance where certaininformation can be found in the application, but are not intended to bethe only source in the application where information on such topic canbe found.

Reference Materials

For other aspects of the polynucleotides, reference is made to standardtextbooks of molecular biology. See, e.g., Hames et al., PolynucleotideHybridization, IL Press, 1985; Davis et al., Basic Methods in MolecularBiology, Elsevir Sciences Publishing, Inc., New York, 1986; Sambrook etal., Molecular Cloning, CSH Press, 1989; Howe, Gene Cloning andManipulation, Cambridge University Press, 1995; Ausubel et al., CurrentProtocols in Molecular Biology John Wiley & Sons, Inc., 1994-1998.

Without further elaboration, it is believed that one skilled in the artcan, using the preceding description, utilize the present invention toits fullest extent. The following preferred specific embodiments are,therefore, to be construed as merely illustrative, and not limitative ofthe remainder of the disclosure in any way whatsoever. The entiredisclosure of all applications, patents and publications, cited aboveare hereby incorporated by reference in their entirety, including, U.S.Provisional Application Nos. 60/331,042 which was filed Nov. 7, 2001,60/331,041 which was filed Nov. 7, 2001, 60/340,251 which was filed Dec.18, 2001, and 60/344,791 which was filed Jan. 7, 2002. TABLE 1 Type ofClone seq Genomic Seq. Clone name Polymorphism (Pos/nt) (Accn#, nt)Pc219 substitution  941, G NT_010736, T substitution  997, A NT_010736,G substitution 1189, G NT_010736, T Pc444 substitution 1024, ANT_008043, G substitution 1028, T NT_008043, C substitution 1085, TNT_008043, C Insertion 1228-1229, CT NT_008043, ** substitution 1277, ANT_008043, G substitution 1396, C NT_008043, G substitution 1480, CNT_008043, T substitution 1498, T NT_008043, A substitution 1924, CNT_008043, T Deletion 1950-1951, ** NT_008043, AC substitution 2346, ANT_008043, T substitution 2481, C NT_008043, G substitution 2973, GNT_008043, A substitution 3861, G NT_008043, A Pc011 Insertion  679, AAC007563, * Insertion 3977, T AC007563, * Pc287 substitution 2287, AAL137878, G substitution 2840, C AL137878, T substitution 2885, AAL137878, G Pc382 substitution  733, G AC000119, A substitution 1479, AAC000119, G substitution 2969, C AC000119, T substitution 3780, CAC000119, T substitution 5959, T AC000119, C

1. A method of determining the presence of prostate cancer cells in asample comprising nucleic acid, comprising: contacting said sample witha polynucleotide probe under conditions effective for said probe tohybridize specifically to a target nucleic acid in said sample,detecting hybridization between said probe and target nucleic acid, anddetermining by said hybridization whether said target nucleic acid isdifferentially-regulated in said sample, whereby the presence of adifferentially-regulated target nucleic acid indicates that said samplecomprises cancer cells, wherein said probe is a polynucleotide of claim29 which is selected from SEQ ID NO 1, 3, 5, 7, 9, 11, 13, 15, 17, 19,21, 23, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 44, 46, or 48, apolynucleotide having 95% sequence identity or more to a sequence setforth in SEQ ID NO 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 24, 26,28, 30, 32, 34, 36, 38, 40, 42, 44, 46, or 48, effective specificfragments thereof, or complements thereto.
 2. A method of claim 1,wherein said determining comprises: comparing the amount ofhybridization in said sample with the amount of hybridization of saidprobe in a second sample comprising normal prostate.
 3. A method ofclaim 1, wherein said determining comprises: comparing the amount ofhybridization in said sample with the amount of hybridization between asecond probe and its corresponding second target nucleic acid in saidsample.
 4. A method of claim 1, wherein said probe is a contiguoussequence of at least 14 nucleotides selected from the sequence set forthin SEQ ID NO 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 24, 26, 28, 30,32, 34, 36, 38, 40, 42, 44, 46, or 48, or a complement thereto.
 5. Amethod of claim 1, wherein said detecting is performed by Northern blotanalysis, polymerase chain reaction (PCR), reverse transcriptase PCR,RACE PCR, or in situ hybridization.
 6. A method of claim 1, wherein saidsample is blood, stool, urine, or prostate tissue.
 7. A method fordiagnosing a prostate cancer in a sample comprising prostate tissue,comprising: determining the number of target genes which aredifferentially-regulated in said sample, wherein said target genes areselected from SEQ ID NO 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 24,26, 28, 30, 32, 34, 36, 38, 40, 42, 44, 46, or 48 of claim 29, or, agene represented by a sequence having 95% sequence identity or more to asequence selected from SEQ ID NO 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21,23, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 44, 46, or 48, wherein saidgenes are differentially-regulated in prostate cancer, and whereby saidnumber is indicative of the probability that said sample comprisesprostate cancer.
 8. A method of claim 7, wherein said determining isperformed by Northern blot analysis, polymerase chain reaction (PCR),reverse transcriptase PCR, RACE PCR, or in situ hybridization using apolynucleotide probe which is SEQ ID NO 1, 3, 5, 7, 9, 11, 13, 15, 17,19, 21, 23, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 44, 46, or 48, apolynucleotide having 95% sequence identity or more to a sequence setforth in SEQ ID NO 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 24, 26,28, 30, 32, 34, 36, 38, 40, 42, 44, 46, or 48, effective specificfragments thereof, or complements thereto.
 9. A method of claim 7,wherein said determining is performed by: contacting said sample with apolynucleotide probe under conditions effective for said probe tohybridize specifically to a target nucleic acid in said sample, anddetecting the amount of hybridization between said probe and targetnucleic acid, and comparing the amount of hybridization in said samplewith the amount of hybridization of said probe in a second samplecomprising normal prostate tissue.
 10. A method of claim 7, wherein saiddetermining is performed by: contacting said sample with apolynucleotide probe under conditions effective for said probe tohybridize specifically to a target nucleic acid in said sample, anddetecting the amount of hybridization between said probe and targetnucleic acid, and comparing the amount of hybridization in said samplewith the amount of hybridization between a second probe and itscorresponding second target nucleic acid in said sample.
 11. A method ofclaim 7, wherein said probe is a contiguous sequence of at least 14nucleotides selected from a sequence set forth in SEQ ID NO 1, 3, 5, 7,9, 11, 13, 15, 17, 19, 21, 23, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42,44, 46, or 48, or a complement thereto. 12-13. (canceled)
 14. A methodfor identifying agents that modulate the expression of targetpolynucleotides differentially-regulated in prostate cancer cells,comprising, contacting a prostate cell population with a test agentunder conditions effective for said test agent to modulate theexpression of a target polynucleotide in said cell population, anddetermining whether said test agent modulates said target polynucleotideexpression, wherein said target polynucleotide is SEQ ID NO 1, 3, 5, 7,9, 11, 13, 15, 17, 19, 21, 23, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42,44, 46, or 48, a polynucleotide having 95% sequence identity or more toa sequence set forth in SEQ ID NO 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21,23, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 44, 46, or 48 of claim 29,effective specific fragments thereof, or complements thereto, and saidpolynucleotide is differentially-regulated in a prostate cancer.
 15. Amethod of claim 14, wherein said agent is an antisense polynucleotide toa target polynucleotide sequence selected from SEQ ID NO 1, 3, 5, 7, 9,11, 13, 15, 17, 19, 21, 23, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 44,46, or 48 and which is effective to inhibit translation of said targetpolynucleotide.
 16. A method for identifying agents that modulate abiological activity of a polypeptide differentially-regulated inprostate cancer cells, comprising, contacting a polypeptidedifferentially-regulated in prostate cancer cells with a test agentunder conditions effective for said test agent to modulate a biologicalactivity of said polypeptide, and determining whether said test agentmodulates said biological activity, wherein said polypeptide is codedfor by a polynucleotide selected from SEQ ID NO 1, 3, 5, 7, 9, 11, 13,15, 17, 19, 21, 23, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 44, 46, or48 of claim 29, a polynucleotide having 95% sequence identity or more toa sequence set forth in SEQ ID NO 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21,23, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 44, 46, or 48, effectivespecific fragments thereof, or complements thereto, and saidpolynucleotide is differentially-regulated in a prostate cancer. 17-18.(canceled)
 19. A method of diagnosing a prostate cancer comprising:assessing the expression of at least one gene selected from SEQ ID NO 1,3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 24, 26, 28, 30, 32, 34, 36, 38,40, 42, 44, 46, or 48 of claim 29, wherein said gene isdifferentially-regulated in said cancer.
 20. A method of claim 19,wherein assessing is: measuring mRNA expression levels of said ormeasuring the expression levels of polypeptide coded for by said gene.21. A method of claim 19, further comprising: comparing said expressionto the expression of said gene of a known normal tissue.
 22. (canceled)23. A method of retrieving prostate cancer differentially-regulated genesequences from a computer-readable medium, comprising: selecting a geneexpression profile that specifies that said gene isdifferentially-regulated in a prostate cancer, and retrieving prostatecancer differentially-regulated gene sequences, where the gene sequencesconsist of SEQ ID NO 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 24, 26,28, 30, 32, 34, 36, 38, 40, 42, 44, 46, or 48 of claim 29, apolynucleotide having 95% sequence identity or more to a sequence setforth in SEQ ID NO 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 24, 26,28, 30, 32, 34, 36, 38, 40, 42, 44, 46, or 48, effective specificfragments thereof, or complements thereto.
 24. (canceled)
 25. Acomposition for detecting a differentially-regulated prostate cancergene, comprising: a pair of polynucleotide primers, each pair comprisinga forward and reverse primer which are effective for specificallyamplifying a polynucleotide which is SEQ ID NO 1, 3, 5, 7, 9, 11, 13,15, 17, 19, 21, 23, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 44, 46, or48 of claim 29 or a complement thereto, wherein, each primer consists of8-100 nucleotides.
 26. An ordered array of polynucleotide probes fordetecting the expression of differentially-regulated prostate cancergenes in a sample, comprising: polynucleotide probes associated with asolid support, wherein each probe is specific for a differentdifferentially-regulated prostate cancer gene, and the probes comprise apolynucleotide which is selected from SEQ ID NO 1, 3, 5, 7, 9, 11, 13,15, 17, 19, 21, 23, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 44, 46, or48 of claim 29, or a complement thereto.
 27. A computer-readable storagemedium, consisting essentially of, differentially up-regulated cancerprostate genes which are selected from SEQ ID NO 1-24, a polynucleotidehaving 95% sequence identity or more to a sequence set forth in SEQ IDNOS 1-24, effective specific fragments thereof, or complements thereto,and said polynucleotide is up-regulated in said prostate cancer. 28.(canceled)
 29. An isolated polynucleotide comprising, a polynucleotidesequence set forth in SEQ ID NO 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21,23, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 44, 46, or 48, or acomplement thereto.
 30. An isolated polypeptide comprising, the aminoacid sequence set forth in SEQ ID NO 1-24. 31-32. (canceled)
 33. Anantibody which is specific for a polypeptide coded for by a geneselected from SEQ ID NO 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 24,26, 28, 30, 32, 34, 36, 38, 40, 42, 44, 46, or 48 of claim 29.